Nucleic Acids Symp Ser, 1980, (7), 387 - 95 Applications of synthetic oligodeoxyribonucleotides to problems in molecular biology; Smith M; The applications of synthetic oligodeoxyribonucleotides to problems in molecular biology described in this article are those where the oligodeoxyribonucleotide is a probe for a specific region of a nucleic acid . This includes the isolation of the iso-1-cytochrome c gene of yeast; the sequence determination of RNAs and DNAs including regions of double-stranded DNA; the introduction of defined site-specific point mutations into bacteriophage OX174 and in the in vitro selection of mutant DNA from a mixture with wild-type DNA. Mol Gen Genet, 1980, 179(1), 55 - 61 Studies on the E . coli groNB (nusB) gene which affects bacteriophage lambda N gene function; Georgopoulos CP et al.; Escherichia coli mutants, called groNB, which block the growth of bacteriophage lambda at the level of action of the gene N product, have been isolated as survivors at 42 degrees C of bacteria carrying a) the defective prophage lambda bio11 i lambda cI857 delta H1 or b) the pcR1 plasmid containing the EcoRI immunity fragment of phage lambda cI857 . In addition, groNB bacterial mutants have been isolated at 37 degrees C, as large colony formers in the presence of lambda i lambda cI h434, lambda i lambda cI h lambda, and lambda i lambda cI h80 phage . The groNB locus is located at 9 minute of the E . coli genetic map with the order of the neighboring loci being proC tsx groNB purE . Most groNB mutations isolated at 42 degrees C were found to interfere in addition with bacterial growth at low temperatures, since (a) the GroNB phenotypes of lambda growth inhibition and bacterial cold sensitivity cannot be separated by P1 transduction, and (b) some cold resistant revertants simultaneously become Gro+ for lambda growth . Lambda transducing phages carrying the groNB+ bacterial gene have been isolated . GroNB mutant bacteria lysogenized by the transducing phage acquire the Gro+ phenotype and simultaneously the cold resistant phenotype, suggesting that the groNB mutations are recessive to the wild-type gene. Proc Natl Acad Sci U S A, 1980 Jan, 77(1), 513 - 7 Chromosomal mutations of Escherichia coli that alter expression of conjugative plasmid functions; McEwen J et al.; We have identified two chromosomal genes of Escherichia coli K12 that are required for the expression of conjugative plasmid functions in the presence of normal plasmid DNA . Hfr cells with mutations in both of these genes are resistant to donor-specific bacteriophage and defective as conjugal donors . These characteristics can be attributed to the inability of mutant Hfr cells to elaborate F-pili, surface organelles required both for conjugal donor ability and for sensitivity to donor-specific bacteriophages . Mutant cells are also defective in surface exclusion, the property of donor cells to act as poor conjugal recipients . This defect can be attributed in part to a reduction in the amount of the F-plasmid traT gene product in the outer membrane of mutant cells; this protein is one of two plasmid gene products required for the full expression of surface exclusion . We have designated the chromosomal genes identified by these mutations as cpxA and cpxB; the mnemonic cpx signifying conjugative plasmid expression. Biochemistry, 1979 Dec 25, 18(26), 5751 - 6 Selective repression of transcription by base sequence specific synthetic polymers; Kosturko LD et al.; We report the effect of novel synthetic polymers on deoxyribonucleic acid (DNA) directed ribonucleic acid (RNA) synthesis in vitro . Polymers contained base-selective monomers, including a GC-specific phenazine derivative and an AT-specific triphenylmethane dye . Radical chain polymerization was carried out in aqueous solution by using monomers bound to a template DNA, which was obtained from either lambda or T7 bacteriophage . Polymers were isolated and reannealed with DNA samples, including competitive mixtures of T7 and lambda DNAs . We measured transcription from DNA-polymer complexes by using Escherichia coli RNA polymerase and determined not only the reduction in total transcription levels but also the relative inhibition of lambda- or T7-specific transcription by using a hybridization assay . The results show that micromolar concentrations of individual dyes are sufficient to cause substantial inhibition of transcription when the dyes are incorporated into polymers . More significantly, a number of the polymers inhibited more strongly transcription from the DNA which had served as template for polymer synthesis than from the DNA present as competitor in the annealing process . We conclude that template synthesis of DNA-binding polymers can lead to preferential inhibition of function of the original template . The apparent relative affinity of polymer for competing DNAs can be altered by at least an order of magnitude depending on which DNA was used as the synthesis template . The results offer a new approach to improving the specificity of DNA-binding drugs. J Biol Chem, 1979 Dec 25, 254(24), 12642 - 6 Bacteriophage fd gene II-protein . II . Specific cleavage and relaxation of supercoiled RF from filamentous phages; Meyer TF et al.; Bacteriophage fd gene II-protein was characterized as an endonuclease which specifically nicked supercoiled replicative form (RF) of filamentous phages in the viral strand . No other supercoiled DNAs tested were attacked by the enzyme, nor were doubly closed fd RF in the relaxed state nor phage fd single strands . Maximal activity was found at pH 8.5 and 80 mM KCl using fd RFI of physiological superhelicity . Mg2+, but no other cofactor, was required for the cleavage reaction . A sealing activity was found to be associated with the enzyme . At a higher concentration of Mg2+ up to 40% of the reaction products were found as doubly closed relaxed fd RF . The protein was not found to be tightly attached to the cleaved strand. J Biol Chem, 1979 Dec 25, 254(24), 12615 - 28 dnaG (primase)-dependent origins of DNA replication . Nucleotide sequences of the negative strand initiation sites of bacteriophages St-1, phi K, and alpha 3; Sims J et al.; The simplest known origins of DNA replication occur in the single-stranded bacteriophages . In one set of phages, negative strand synthesis is initiated by a single protein, the product of the Escherichia coli replication gene dnaG . Evidently, in these phages--G4, St-1, phi K, and alpha 3--the origin for negative strand synthesis consists of a nucleic acid element capable of direct recognition by the dnaG priming protein . We have located and sequenced the origins of negative strand synthesis in St-1, phi K, and alpha 3, and compared them with the origin sequence previously determined for G4 . In each case, the point at which the negative strand is initiated can be identified at the nucleotide level . The data lead to the following conclusions: 1 . In all four phages, the negative strand initiation site occurs within an intercistronic region of approximately 135 bases . While in G4, the origin lies between genes specifying the viral coat proteins F and G, the origin is shifted in St-1, phi K, and alpha 3 to a position between coat protein genes G and H . 2 . Extensive nucleotide conservation exists at the negative strand origin, but does not extend into the adjacent coding regions . The conserved origin DNA occurs in two regions, 42 and 45 bases long, which are separated by 13 bases of divergent sequence . 3 . Correlated with the two stretches of conserved nucleotide sequence are two regions of potential secondary structure . The start point of negative strand synthesis lies just prior to one of these hairpins . Similarities in both primary sequence and secondary structure can be found between the negative strand origins of G4, St-1, phi K, and alpha 3 and the general origin regions of bacteriophage lambda and of E . coli. J Biol Chem, 1979 Dec 25, 254(24), 12419 - 26 Mutants of Escherichia coli that do not contain 1,4-diaminobutane (putrescine) or spermidine; Hafner EW et al.; Strains of Escherichia coli K12 have been constructed which do not contain any of the polyamines normally present in a wild type strain, namely, 1,4-diaminobutane (putrescine) and spermidine . This phenotype arises as a consequence of the assembly into these strains of deletion mutations in speA (arginine decarboxylase), speB (agmatine ureohydrolase), speC (ornithine decarboxylase), and speD (adenosylmethionine decarboxylase) . The polyamine-deficient strains grow indefinitely in the absence of polyamines but with a growth rate one-third of that found in the presence of polyamines . These strains can act as hosts for bacteriophages T4, T7, and f2, although the latter phage is poorly adsorbed; they can also maintain F' factors, ColE1 and P1 plasmids, and lysogeny by bacteriophage lambda . In contrast, the production of bacteriophage lambda in the absence of polyamines is strikingly decreased (greater than 99%) either after infection of a nonlysogen or after induction of a lysogen . A polyamine-deficient Hfr strain can transfer its chromosome to a recipient at a normal rate, but the number of recombinants observed in a cross is decreased approximately 300-fold . No such effect is observed when only the F- recipient strain in a cross is polyamine deficient. Science, 1979 Dec 21, 206(4425), 1419 - 21 Visualization of polymercurimethane-labeled for bacteriophage in the scanning transmission electron microscope; Lipka JJ et al.; Each of the 2700 coat proteins of fd bacteriophage was labeled with tetrakis(acetoxymercuri)methane (TAMM) or aquoglycylmethionineplatinum(II) . The TAMM-labeled specimens reveal striking bright spots in the scanning transmission electron microscope which arise from clustering . Measurements of mass show increases consistent with the addition of four mercury atoms or one platinum atom, respectively, to each coat protein. Nucleic Acids Res, 1979 Dec 20, 7(8), 2457 - 67 Nucleosome dissociation and transfer in concentrated salt solutions; Stacks PC et al.; We have examined the dissociation of nucleosomes into histones and free, 4.5S DNA over a range of sodium chloride concentrations between 0.25 and 1 M . We have also studied this dissociation as a function of nucleosome concentration at two salt concentrations, 0.8 M and 0.9 M . In addition, we have measured the kinetics of transfer of histone cores from nucleosomes onto recipient bacteriophage T7 DNA in 0.6, 0.7 and 0.8 M NaCl solutions . Although the mechanism of nucleosome transfer is unknown the data presented here are consistent with either a reversible dissociation of the nucleosome or DNA strand displacement by another DNA. Nucleic Acids Res, 1979 Dec 20, 7(8), 2255 - 73 Studies on the binding of lambda Int protein to attachment site DNA: identification of a tight-binding site in the P' region; Davies RW et al.; We have used three approaches to studying the interaction of lambda Int protein with bacteriophage attachment site DNA, POP': location of binding sites by retention of DNA fragments in a filter binding assay, reconstruction of a binding site by DNA synthesis and protection of a binding site from an exonuclease . Retention of restriction fragments on nitrocellulose filters in the presence of Int protein was used to locate binding sites . A high affinity binding site lies in P' between base pairs -6 and +173 from the center of the common core sequence, and low affinity sites are found in the 200 base pair region left of position -6 . Reconstruction of the high affinity binding site region from the right using primed DNA synthesis and testing for filter binding in the presence of Int protein shows that sequences sufficient for tight binding of Int protein lie to the right of position +66 . When attachment site DNA is protected by bound Int protein against digestion by exonuclease III, four Int dependent protection bands are seen in positions +58, +68, +79 and +88 . This can be interpreted either as showing that four Int protein monomers bind to the high affinity region in series, or as evidence for wrapping of the DNA around Int protein, leading to structural changes resembling those occurring to DNA in nucleosomes. Nucleic Acids Res, 1979 Dec 20, 7(8), 2177 - 88 Cleavage of single-stranded DNA by the A and A* proteins of bacteriophage phi X174; Langeveld SA et al.; The purified A protein and A* protein of bacteriophage phi X174 have been tested for endonuclease activity on single stranded viral phi X174 DNA . The A protein (55.000 daltons) nicks single-stranded DNA in the same way and at the same place as it does superhelical RFI DNA, at the origin of DNA replication . The A* protein (37.000 daltons) can cleave the single-stranded viral DNA at many different sites . It has however a strong preference for the origin of replication . Both proteins generate 3'OH ends and blocked 5' termini at the nick site. Eur J Biochem, 1979 Dec 17, 102(2), 589 - 94 Crystallization of bacteriophage MS2; Min Jou W et al.; Crystals of bacteriophage MS2 have been obtained by slowly cooling a 1% virus solution from 23 degrees C to 0 degrees C in the presence of poly(ethylene glycol) 6000 . The crystals were colorless, needle-like, anisotropic and very fragile . Electron microscopic observation of the crystals revealed a two-dimensional lattice of particles with RNA phage morphology and dimensions . Preliminary X-ray examination of the crystals confirmed their viral nature. Eur J Biochem, 1979 Dec 17, 102(2), 595 - 604 Secondary structure of the 5' end of bacteriophage MS2 RNA Methoxyamine and kethoxal modification; Iserentant D et al.; To refine the secondary structure model of the 5' end of the bacteriophage MS2 genome, 32P-labeled MS2 RNA was partially digested with T1 RNase or with Cm-RNase and the 5'-end fragment was isolated, renatured and submitted to treatment with methoxyamine or kethoxal . The resulting modified RNA was digested with T1 RNase and the products were separated by minifingerprinting . Methoxyamine-induced modification of exposed cytidines was detected by differential mobility of modified oligonucleotides, while kethoxal-induced alteration of exposed guanosines was monitored by resistance to T1 ribonuclease digestion . The positions of the modified residues are discussed in terms of an improved secondary structure model proposed for the 5' end of the viral RNA . The structure itself is discussed in relation to sequence conservation and biological function. Gene, 1979 Dec, 8(1), 53 - 68 Cloning of bacteriophage T5 DNA fragments in plasmid pBR322 and bacteriophage lambda gtWES; Brunel F et al.; Bacteriophage T5 was digested with the restriction endonucleases HindIII and EcoRI and the resulting fragments were inserted into the plasmid pBR322 and the bacteriophage lambda gtWES as vectors . Approx . 15% of the phage genome was recovered in recombinant clones . The recombinants were characterized by restriction analysis, DNA/DNA hybridization employing Southern blots, and ability to complement or recombine with amber mutants of T5 . The results obtained allow revisions of the physical map of the T5 genome and partial correlation of the physical map with the genetic map. Cell, 1979 Dec, 18(4), 1273 - 83 The linkage arrangement of four rabbit beta-like globin genes; Lacy E et al.; Four different regions of rabbit beta-like globin gene sequences designated beta 1, beta 2, beta 3 and beta 4 were identified in a set of clones isolated from a bacteriophage lambda library of chromosomal DNA fragments (Maniatis et al., 1978) . Restriction mapping and blot hybridization (Southern, 1975) studies indicate that a subset of these clones containing beta 1 and beta 2 hybridizes to an adult beta-globin cDNA clone (Maniatis et al., 1976) more efficiently than to a human gamma-globin cDNA clone (Wilson et al., 1978), while another subset containing beta 3 and beta 4 displays the converse hybridization specificity . beta 1 was identified as the adult beta-globin gene, while beta 2, beta 3 and beta 4 have not been identified with any known rabbit globin polypeptides . Cross-hybridization and transcriptional orientation experiments indicate that the set of beta-like gene clones contains overlapping restriction fragments encompassing 44 kb of rabbit chromosomal DNA . In addition, all four genes have the same transcriptional orientation and are arranged in the order 5'-beta 4-beta 3-beta 2-beta 1-3'. J Virol, 1979 Dec, 32(3), 905 - 16 regA protein of bacteriophage T4D: identification, schedule of synthesis, and autogenous regulation; Cardillo TS et al.; Proteins labeled with 14C-amino acids after infection of Escherichia coli B by T4 phage were examined by electrophoresis in the presence of sodium dodecyl sulfate . Four regA mutants (regA1, regA8, regA11, and regA15) failed to make a protein having a molecular weight of about 12,000, whereas mutant regA9 did make such a protein; regA15 produced a new, apparently smaller protein that was presumably a nonsense fragment, whereas regA11 produced a new, apparently larger protein . We conclude that the 12,000-dalton protein was the product of the regA gene . The molecular weight assignment rested primarily on our finding that the regA protein had the same mobility as the T4 gene 33 protein, which we identified by electrophoresis of whole-cell extracts of E . coli B infected with a gene 33 mutant, amE1120 . Synthesis of wild-type regA protein occurred from about 3 to 11 min after infection at 37 degrees C in the DNA+ state and extended to about 20 min in the DNA- state . However, synthesis of the altered regA proteins of regA9, regA11, and regA15 occurred at a higher rate and for a much longer period in both the DNA+ and DNA- states; thus, the regA gene is autogenously regulated . At 30 degrees C, both regA9 and regA11 exhibited partial regA function by eventually shutting off the synthesis of many T4 early proteins; the specificity of this shutoff differed between these two mutants . We also obtained evidence that the regA protein is not Stevens's "polypeptide 3." As a technical point, we found that, when quantitating acid-precipitable radioactivity in protein samples containing sodium dodecyl sulfate, it was necessary to use 15 to 20% trichloroacetic acid; use of 5% acid, e.g., resulted in loss of over half of the labeled protein. Proc Natl Acad Sci U S A, 1979 Dec, 76(12), 6314 - 8 Molecular cloning of Moloney murine sarcoma virus: arrangement of virus-related sequences within the normal mouse genome; Tronick SR et al.; The unintegrated circular DNA form of Moloney murine sarcoma virus (MSV) has been cloned in bacteriophage lambda . Discrete deletions in the viral genome were shown to occur during propagation of recombinant phage in Escherichia coli . Heteroduplex and restriction enzyme analyses indicated the deletion of tandemly repeated sequences within certain of the cloned MSV DNA inserts . Cloned MSV DNA was used to prepare a probe composed of its acquired cellular (src) sequences, shown previously to be necessary for MSV transformation . Analysis of EcoRI digests of normal mouse cellular DNA revealed the presence of a single 14-kilobase-pair fragment containing these sequences which lacked contiguity with endogenous type C helper viral information of the same cells . Thus, the sarcoma virus-specific sequences of MSV are represented within the normal mouse genome in a manner analogous to that of a cellular gene. Mutat Res, 1979 Dec, 63(2), 273 - 89 Biological and chemical effects of mustard gas in yeast; Kircher M et al.; Mustard gas induces inactivation and mutation in yeast . Both effects are dose-proportional, indicating single-hit events . Induction of both effects is influenced by the cell's capacity for DNA dark-repair, whereby the probability of reversion is highest in repair-proficient cells . Binding of mustard gas to cells and probably to DNA is independent of DNA-repair systems . The number of inter-strand cross-links, as determined by assaying for renaturability of alkalidenatured DNA, increases in a dose-proportional manner . At 37% survival an excision-deficient strain contains 55 inter-strand cross-links . Chromatographic analysis yields several alkylation products of DNA . Their relative frequencies resemble the values reported for E . coli and bacteriophage T7. J Bacteriol, 1979 Dec, 140(3), 1071 - 80 Interaction of bacteriophage T4 with reconstituted cell envelopes of Escherichia coli K-12; Furukawa H et al.; The interaction with bacteriophage T4 of the cell surface of Escherichia coli K-12 reconstituted from outer membrane protein O-8, lipopolysaccharide, and the lipoprotein-bearing peptidoglycan sacculus was studied . The reconstituted cell surface was active as a receptor for the phage, resulting in the contraction of the tail sheath, a morphological change in the base plate which was accompanied by the extension of short tail pins down to the cell surface and the penetration of the needle through the cell surface . However, the ejection of phage deoxyribonucleic acid did not take place . Both O-8 and lipopolysaccharide were essential for the interaction . In the reconstitution, the wild-type lipopolysaccharide could not be replaced by either heptoseless lipopolysaccharide or lipid A . The lipoprotein-bearing peptidoglycan sacculus was also found to be an active component for the phage adsorption . The sacculus most likely functioned as a basal framework on which O-8 and lipopolysaccharide assembled to form a flat sheet which is large enough to interact with individual distal ends of long tail fibers of a single phage particle. J Virol, 1979 Dec, 32(3), 917 - 24 Inhibition of initiation of bacteriophage T4 DNA replication by perturbation of Escherichia coli host membrane composition; Huang WM; 3-Decynoyl-N-acetylcysteamine (3-decynoyl-NAC) is an analog which specifically causes the immediate cessation of the biosynthesis of unsaturated fatty acids in Escherichia coli, whereas the synthesis of saturated fatty acids is actually stimulated . As a result, the cell membrane accumulates saturated fatty acids in its phospholipid . Addition of the inhibitor at the time of infection of E . coli by T4 phage had no effect on normal phage replication and development, implying that the synthesis of unsaturated fatty acids per se has little effect on T4 DNA replication . However, if the integrity and composition of the bacterial membrane was grossly perturbed by first treating the cells with the inhibitor for 60 min before infection, the proper initiation and the attainment of a rapid rate of T4 DNA synthesis were not observed . Under these conditions, a full complement of T4 early proteins was synthesized . The membrane associability of the known DNA delay proteins induced by wild-type T4 phage in the treated cells resembled that expected of a culture of untreated cells infected with a DNA delay mutant . When any one of three DNA delay mutants was used to infect 3-decynoyl-NAC-treated cells, T4 DNA replication was aborted . These findings suggest that some kind of specific interactions among the initiation proteins defined by the DNA delay mutants and the bacterial membrane may be necessary to facilitate the normal initiation and rapid rate of T4 DNA replication . A model for the involvement of the three different initiation proteins and the subsequent attainment of rapid DNA synthesis is discussed. Proc Natl Acad Sci U S A, 1979 Dec, 76(12), 6240 - 4 Cell-cycle-associated rearrangement of inverted repeat DNA sequences; Nisen P et al.; Inverted repeat DNA sequences of Caulobacter crescentus have been isolated, characterized, and cloned in a bacteriophage lambda vector . Both whole populations and individual clones of these sequences were hybridized to restriction endonuclease-generated fragments of chromosomal DNA isolated from cells that were in different stages of the cell cycle . Some inverted repeat DNA sequences were observed to hybridize to different regions of the chromosomal DNA isolated from the morphologically and biochemically distinct swarmer cell and stalked cell populations . These results suggest that the inverted repeat sequences have the capacity to rearrange and thus be located at different sites on the genomes of the different cell types. Proc Natl Acad Sci U S A, 1979 Dec, 76(12), 6137 - 41 Origins of phage T4 DNA replication as revealed by hybridization to cloned genes; Halpern ME et al.; {3H}Thymidine-labeled progeny DNA was isolated after infection of Escherichia coli with two different bacteriophage T4 mutants . These strands were isolated shortly after the initiation of DNA replication and hybridized to 15 different (EcoRI) T4 restriction fragments cloned in plasmid vectors . Uniformly labeled T4 {32P}DNA extracted from phage particles was cohybridized as a normalizing reference . The results obtained lead to the conclusions that, among the loci tested, initiation occurs predominantly in the area of genes 50-5 and less prominently in the area of genes 25-29 . However, our data do not support the idea of initiation in the area of genes 40-43 . In contrast, this area displays the least replication among the genes tested. Gene, 1979 Dec, 8(1), 69 - 80 A new host-vector system allowing selection for foreign DNA inserts in bacteriophage lambda gtWES; Davison J et al.; An improved vector (lambda gtWES.T5-622) for EcoRI fragments has been derived from EK2 vector lambda gtWES.lambdaB' by replacing the lambda B fragment with two identical 1.1 Md fragments from the pre-early region of bacteriophage T5 . The new vector has two advantages which facilitate elimination of parental-type recombinants in an in vitro recombination experiment . Firstly, the 1.1 Md insert is too small to be re-inserted into lambda gtWES in a single copy . Secondly the 1.1 Md T5 fragment carries T5 gene A3 which prevents growth of phage retaining this fragment when the Excherichia coli host carries plasmid ColIb . Thus, essentially all plaques are due to phage with donor DNA inserts and are free of T5 DNA fragments . The size usually given as the theoretical minimum size for insertion into the lambda gt series of vectors is 0.66 Md . We have shown that this size is an underestimate and that the lower limit is about 1.6 Md . A precise estimate is difficult since there is strong selection, among phage having small inserts, for those which have acquired additional genetic material by duplication of the lambda DNA. Proc Natl Acad Sci U S A, 1979 Dec, 76(12), 6066 - 70 DNA from recombinogenic lambda bacteriophages generated by arl mutant of Escherichia coli is cleaved by single-strand-specific endonuclease S1; Hays JB et al.; When propagated on arl strains (a subclass of Escherichia coli hyper-rec mutants), lambda "Red-" duplication phages accumulated an enhanced potential for recombination . The physical properties of the recombinogenic phages thus obtained ("Arl-" phages) were similar to those of phages grown on arl+ bacteria . However, Arl- phage DNA was cleaved by endonuclease S1 under conditions such that the nuclease is specific for single-stranded DNA;DNA from control phages was S1-resistant . The number of S1 sites (defined by the apparent decrease in single-strand molecular weight) reached a maximum (seven to nine sites per strand of lambda DNA) after five or six rounds of growth on arl bacteria . Similarly, the recombinogenicity of Arl- phages reached a limiting value (recombination frequency, 15%) that was 5 times that of Arl+ phages . Recombinogenicity and S1 susceptibility were accumulated concomitantly during growth on arl+ bacteria . If all increased recombination occurred at the S1 sites, then these regions (about 40 bases each) were about 300 times as recombinogenic as normal DNA regions of the same size, and 1.5 times as recombinogenic as UV-induced lesions . Chromosomal DNA and plasmid DNA (pBR322) from arl cells were more susceptible to nuclease S1 than was DNA from arl+ bacteria . Analysis of the cleavage products suggests that the S1 sites on Arl- lambda phage DNA are located randomly. Cell, 1979 Dec, 18(4), 1145 - 51 Specificity of the bacteriophage lambda N gene product (pN): nut sequences are necessary and sufficient for antitermination by pN; de Crombrugghe B et al.; We have cloned the nutR site together with the tR1 site of bacteriophage lambda in the E . coli galactose operon to examine whether the lambda promoter sequences PR and PL are involved in the recognition specificity of the lambda N gene product (pN) . We first constructed a derivative of plasmid pBR322 in which the expression of the tetracycline genes (tet) is controlled by the gal promoter (Pgal) . This new plasmid contains a unique Hind III site between Pgal and tet into which the nutR and tR1 sites were introduced . The order of the relevant genetic markers in this second plasmid is Pgal-nutR-tR1-tet . Cells transformed with this plasmid express tet only if pN is provided and if the plasmid contains an intact gal promoter . Our data suggest that transcription which originates at Pgal is modified by pN at nutR, enabling it to pass through tR1 into tet . We conclude that promoters do not play a specific role in pN recognition and that nut sequences are both necessary and sufficient for pN action. J Virol, 1979 Dec, 32(3), 951 - 7 Characterization of a bacteriophage related to R-type pyocins; Kageyama M et al.; A bacteriophage with a contractile tail which shows very similar features to R-type pyocins was isolated and characterized . This phage, named PS17,was purified by DEAE-cellulose chromatography and CsCl density gradient centrifugation . It was a DNA-containing phage, and the density of the purified particles in CsCl was found to be 1.468 . DNA from this phage had a density of 1.720 in CsCl, indicating its guanine plus cytosine content to be 61.2% . The head was polyhedral, 69 nm in diameter, and the tail was 150 nm in length . This phage was neutralized by antiserum preparations against five R-type pyocins, and the antiserum against this phage was active in neutralizing R-type pyocins . The properties of this phage, PS17, were compared with another similar phage, PS3, which was previously reported. Gene, 1979 Dec, 8(1), 107 - 19 The N protein of bacteriophage lambda, defined by its DNA sequence, is highly basic; Franklin NC et al.; Nucleotide sequence has been determined for the restriction fragments and cloned DNA from the pL-N-tL1 region of bacteriophage lambda . A unique reading frame for the N gene is defined by the absence of natural nonsense codons and by the presence of seven nonsense codons generated by mutations in N . This reading frame is initiated at two alternative ATG codons, the second of which is probably the in vivo translation start . Reading is stopped at a single TAG codon . The protein coded is therefore 133 or, more probably, 107 amino acids long, rich in lysine, arginine and proline. J Biol Chem, 1979 Nov 25, 254(22), 11605 - 14 T7-induced DNA polymerase . Characterization of associated exonuclease activities and resolution into biologically active subunits; Adler S et al.; Bacteriophage T7-induced DNA polymerase has been isolated by a procedure suitable for large scale use and which yields near homogeneous enzyme . In addition to previously described DNA polymerase activity and 3' to 5' exonucleolytic activity on single stranded DNA (Grippo, P., and Richardson, C . C . (1971) J . Biol . Chem . 246, 6867-6873), the enzyme also possesses a highly active exonuclease which hydrolyzes duplex substrates with 3' to 5' directionality . The native polymerase has been dissociated using 6 M guanidine HCl and resolved into biologically active subunits: T7 gene 5 protein and Escherichia coli thioredoxin . The phage-specified subunit obtained by this procedure is deficient in DNA polymerase and double strand exonuclease activities, with deficiencies in these activities being apparent at the level of a single turnover . However, it possesses near normal levels of a single strand hydrolytic activity which is identical to that associated with the native polymerase with respect to substrate specificity and suppression of hydrolysis by low levels of deoxyribonucleoside 5'-triphosphates . Thioredoxin forms a molecular complex with the T7 gene 5 protein, and addition of the host protein restores restores DNA polymerase and double strand exonuclease activities to near normal levels. J Biol Chem, 1979 Nov 25, 254(22), 11591 - 7 Deoxyribonucleic acid polymerase of bacteriophage T7 . Purification and properties of the phage-encoded subunit, the gene 5 protein; Hori K et al.; DNA polymerase of bacteriophage T7 is composed of two subunits, the gene 5 protein of the phage and the host-specified thioredoxin . The gene 5 protein has been purified 7400-fold to homogeneity from bacteriophage T7-infected Escherichia coli 7400 trxA cells that lack thioredoxin . The purification procedure has been monitored by using a complementation assay in which thioredoxin interacts with the gene 5 protein to form an active DNA polymerase . The purified gene 5 protein is a single polypeptide having a molecular weight of 87,000 . The gene 5 protein itself has only 1 to 2% of the polymerase activity of T7 DNA polymerase . However, T7 DNA polymerase can be reconstituted by the addition of homogeneous thioredoxin to the gene 5 protein . Optimal reconstitution is obtained when the molar ratio of thioredoxin/gene 5 protein is 150 . Under these conditions, the gene 5 protein attains approximately 80% of the activity of an equal amount of T7 DNA polymerase . The apparent Km for thioredoxin in the reaction to restore DNA polymerase activity is 2.8 x 10(-8) M . The enzymatic properties of the reconstituted enzyme are indistinguishable from those of T7 DNA polymerase synthesized in vivo; the reconstituted polymerase interacts with T7 gene 4 protein to catalyze DNA synthesis on duplex DNA templates. J Biol Chem, 1979 Nov 25, 254(22), 11208 - 17 Spectroscopic analysis of the interaction of Escherichia coli DNA-dependent RNA polymerase with T7 DNA and synthetic polynucleotides; Reisbig RR et al.; We have studied the circular dichroism and ultraviolet difference spectra of T7 bacteriophage DNA and various synthetic polynucleotides upon addition of Escherichia coli RNA polymerase . When RNA polymerase binds nonspecifically to T7 DNA, the CD spectrum shows a decrease in the maximum at 272 but no detectable changes in other regions of the spectrum . This CD change can be compared with those associated with known conformational changes in DNA . Nonspecific binding to RNA polymerase leads to an increase in the winding angle, theta, in T7 DNA . The CD and UV difference spectra for poly{d(A-T)} at 4 degrees C show similar effects . At 25 degrees C, binding of RNA polymerase to poly{d(A-T)} leads to hyperchromicity at 263 nm and to significant changes in CD . These effects are consistent with an opening of the double helix, i.e . melting of a short region of the DNA . The hyperchromicity observed at 263 nm for poly{d(A-T)} is used to determine the number of base pairs disrupted in the binding of RNA polymerase holoenzyme . The melting effect involves about 10 base pairs/RNA polymerase molecule . Changes in the CD of poly(dT) and poly(dA) on binding to RNA polymerase suggest an unstacking of the bases with a change in the backbone conformation . This is further confirmed by the UV difference spectra . We also show direct evidence for differences in the template binding site between holo- and core enzyme, presumably induced by the sigma subunit . By titration of the enzyme with poly(dT) the physical site size of RNA polymerase on single-stranded DNA is approximately equal to 30 bases for both holo- and core enzyme . Titration of poly{d(A-T)} with polymerase places the figure at approximately equal to 28 base pairs for double-stranded DNA. J Biol Chem, 1979 Nov 25, 254(22), 11598 - 604 Deoxyribonucleic acid polymerase of bacteriophage T7 . Characterization of the exonuclease activities of the gene 5 protein and the reconstituted polymerase; Hori K et al.; Homogeneous gene 5 protein of bacteriophage T7, a subunit of T7 DNA polymerase, catalyzes the stepwise hydrolysis of single-stranded DNA in a 3' leads to 5' direction to yield nucleoside 5'-monophosphates . The gene 5 protein itself does not hydrolyze duplex DNA . However, in the presence of Escherichia coli thioredoxin, the host-specified subunit of T7 DNA polymerase, duplex DNA is hydrolyzed in a 3' leads to 5' direction to yield nucleoside 5'-monophosphates . The apparent Km for thioredoxin in the reaction is 4.8 x 10(-8) M, a value similar to that for the apparent Km of thioredoxin in the complementation assay with gene 5 protein to restore T7 DNA polymerase activity . Both exonuclease activities require Mg2+ and a sulfhydryl reagent for optimal activity, and both activities are sensitive to salt concentration . Deoxyribonucleoside 5'-triphosphates inhibit hydrolysis by both exonuclease activities; hydrolysis of single-stranded DNA by the gene 5 protein is inhibited even in the absence of thioredoxin where there is less than 2% active T7 DNA polymerase . E . coli DNA binding protein (helix destabilizing protein) stimulates the hydrolysis of duplex DNA up to 9-fold under conditions where the hydrolysis of the single-stranded DNA is inhibited 4-fold. Biochim Biophys Acta, 1979 Nov 22, 565(1), 51 - 66 The secondary structure of bacteriophage DNA in situ . VIII . The reaction of sodium bisulphite with intraphage cytosine as a probe for studying the DNA-protein interaction; Sklyadneva VB et al.; To obtain data on the viral nucleoprotein a study has been made of the reaction of sodium bisulphite with cytosine in the intraphage DNA of the phage Sd . The CHlO4 hydrolysates of the bisulphite-modified phage Sd have demonstrated a decrease of 18% in the cytosine content and the presence of the products with the properties of cytosyl-amino acids (the main amino acid responsible for the DNA-protein interaction involving cytosine is lysine) . But when prior to hydrolysis the modified phage was disintegrated under mild conditions in 0.1--1 M NaCl solution or Tris-HCl buffer (pH 7), neither the decrease in the cytosine content nor cytosyl-amino acids have been found . An exception is the heating of the phage at 70 degrees C in a medium containing 0.05 M phosphate buffer (pH 7.9--8.5), when an 18% decrease in the cytosine content and subsequent appearance of cytosyl-amino acids have also been observed . The presence of cytosyl-amino acids which are the nucleotide-protein cross-links is confirmed by the results of viscometry, equilibrium centrifugation in cesium sulphate gradient and determinations of the survival percentage . It is suggested that the reaction between bisulphite and cytosine in the phage Sd stops at the stage of the intermediate product C5-C6-dihydro-C6-sulphopyrimidine whose amino group is shielded by interaction with protein (product VII) . This product can exist only under in situ conditions: with disintegration of nucleoprotein (destruction of phage particles or ejection of the DNA) in phosphate-free media the product VII reverts into the initial cytosine . Under the conditions of acid hydrolysis or destruction of phage in the presence of phosphate ions product VII undergoes transamination with cleavage of SO3 and restoration of the C5-C6 double bond producing cytosyl-amino acids . The factors determining the stability of the product VII are discussed. Nucleic Acids Res, 1979 Nov 10, 7(5), 1335 - 41 Nucleotide sequence of a secondary attachment site for bacteriophage lambda on the Escherichia coli chromosome; Csordas-Toth E et al.; The nucleotide sequence of a secondary attachment site for bacteriophage lambda was determined in a region near the rrnB gene at 88 min on the E . coli chromosome . The sequence has a 8 base pair interrupted homology GCT TTTTA to the common core of the primary attachment site (attB) and the corresponding phage sequence (attP) . The site of crossover during integration lies probably between nucleotides -3 and +1 . The flanking regions have no obvious homology to the arms of either attP or attB. J Gen Virol, 1979 Nov, 45(2), 351 - 9 Regulation of imm gene expression in bacteriophage T4-infected cells; Yutsudo M; Two polypeptides (imm-a and imm-b) which are not induced by an immunity mutant T4Dimm2 but by a wild-type strain T4D were identified by sodium dodecyl sulphate (SDS)-polyacrylamide gel electrophoresis . Their mol . wt . were 77 000 and 45 000, respectively . These polypeptides exhibited a similar kinetic pattern of synthesis . Within a few minutes p.i . the primary phage established the system that inhibited imm gene expression of superinfecting phage . This was shown by measuring both the phenotypic expression of immunity and the synthesis of imm gene polypeptides . The expression of two other immediate-early genes, namely genes s and 30, and early gene 33, was not affected by primary infection. J Virol, 1979 Nov, 32(2), 497 - 506 Structural role of the polyglutamate portion of the folate found in T4D bacteriophage baseplate; Kozloff LM et al.; Three types of reagents were used to determine the structural role and location of the polyglutamate portion of the Escherichia coli T4D bacteriophage baseplate dihydropteroyl hexaglutamate . These reagents were examined for their effect in vitro on some of the final steps in phage baseplate morphogenesis . The reagents were (i) a series of oligopeptides composed solely of glutamic acid residues but with various chemical linkages and chain lengths; (ii) a homogeneous preparation of carboxypeptidase G1, an exopeptidase that hydrolyzes carboxyl-terminal glutamates (or aspartates) from simple oligopeptides, including the gamma-glutamyl bonds on folyl polyglutamates as well as the bond between the carboxyl group of the p-aminobenzoyl moiety and the amino group of the first glutamic acid residue of folic acid; and (iii) antisera prepared against a polyglutamate hapten . All three types of reagent markedly inhibited the attachment of the phage long tail fibers to the baseplate . Other steps in baseplate assembly such as the addition of T4D gene 11 or gene 12 products were not affected by any of these reagents . These results indicate that the polyglutamate portion of the folate is located near the attachment site on the bacteriophage baseplate for the long tail fibers. Appl Environ Microbiol, 1979 Nov, 38(5), 840 - 5 Influence of fulvic acid on bacteriophage adsorption and complexation in soil; Bixby RL et al.; The effect of fulvic acid, the major fraction of natural soluble organic matter, on the adsorption of MS2 bacteriophage to soil was investigated in controlled laboratory experiments . Batch experiments together with scanning electron microscopy-energy-dispersive X-ray analysis showed that fulvic acid complexed phage, which prevented its adsorption to soil . Phage strongly adsorbed to soil in the absence of fulvic acid . Phage which was complexed with fulvic acid was not irreversibly inactivated and could become viable under proper conditions, illustrating the importance of assay and elution procedures in the recovery of virus from aqueous solutions. Proc Natl Acad Sci U S A, 1979 Nov, 76(11), 5789 - 93 Isolation and characterization of transducing phage coding for sigma subunit of Escherichia coli RNA polymerase; Gross CA et al.; A transducing phage has been isolated with codes for the sigma subunit of Escherichia coli RNA polymerase . Transducing phage were selected from E . coli shotgun collections of HindIII or Sac I fragments cloned into Charon 25, a new bacteriophage lambda vector that is capble of forming lyosogens at high temperature . Transduction of an E . coli strain carrying a temperature-sensitive mutation in the sigma gene was used for the selection . The positions of restriction sites for Sac I, HindIII, Xho I, Bgl II, and Kpn I in the cloned bacterial DNA segments were determined . Phage containing the HindIII fragment complement both primase (dnaG) and sigma (rpoD) whereas those containing the Sac I fragment complement only sigma . Results of analyses of the proteins made both in vivo after infection of UV-irradiated cells and in vitro in a coupled transcription/translation system suggest that a Sac I site separates the promoter for sigma from the sigma structural gene . The direction of transcription of sigma was determined to be clockwise with respect to the E . coli genetic map. Proc Natl Acad Sci U S A, 1979 Nov, 76(11), 5430 - 4 RNA ligase reaction products in plasmolyzed Escherichia coli cells infected by T4 bacteriophage; David M et al.; Searching for a physiological role of T4 RNA ligase {polyribonucleotide synthetase (ATP); poly(ribonucleotide):poly(ribonucleotide) ligase (AMP-forming), EC 6.5.1.3} activity, we developed an acellular system of plasmolyzed Escherichia coli cells infected by T4 bacteriophage . Upon incubation of this system with {gamma-32P}ATP, 32P was transferred into a large number of polyribonucleotides, mostly up to 300-400 residues long . The bulk of 32P in the product polyribonucleotides was found in 5'-terminal phosphate groups, suggesting that they originated by a phosphorylation reaction catalyzed by the endogenous polynucleotide kinase (EC 2.7.1.78) . Indeed, these products were not seen in an acellular system from uninfected cells, and their amount and complexity increased with the progress of infection . Analysis of the 32P-labeled polyribonucleotide products by gel electrophoresis, either before or after digestion with alkaline phosphatase (EC 3.1.3.1), revealed that a small fraction of the 32P resided in phosphodiester bonds of several tRNA-sized chains . This specific 32P transfer from {gamma-32P}ATP into phosphodiester bonds was apparently catalyzed by successive polynucleotide kinase and RNA ligase reactions . The possible relationship of the 32P transfer to RNA ligase was investigated next by using a system from cells infected with T4 am M69 (an amber mutant deficient in RNA ligase) . Transfer of 32P from {gamma-32P}ATP into phosphodiester bonds was not detected in the am M69 system . However, addition of purified RNA ligase to the am M69 system restored the specific 32P transfer . A system from cells infected with T4 psu-b delta 33 (a deletion mutant lacking the entire tRNA region) sustained the specific 32P transfer into tRNA-sized products, indicating that they were not derived from transcripts of T4 tRNA genes . These data may reflect a role of RNA ligase in posttranscriptional conversion of presumably host polyribonucleotides into novel tRNA species during T4 infection. Mol Gen Genet, 1979 Nov, 176(3), 433 - 8 Cloning of bacteriophage PM2 DNA in Escherichia coli K12; Rempola B et al.; DNA fragments of phage PM2 restricted with Hin dIII endonuclease was cloned in the vector pBR 322 in an Escherichia coli K12 host . The attempt to clone full length PM2 DNA restricted with PstI endonuclease has been unsuccesful . From six randomly chosen recombinant clones DNA was purified and analysed with EcoRI, PstI and Hin dIII endonucleases . The physical map of three chimeric plasmids was unequivocally established . It was shown, that the whole PM2 genome was cloned, although in separate fragments . However, most of the recombinant clones were instable in the absence of selective pressure. J Bacteriol, 1979 Nov, 140(2), 680 - 6 Interaction of bacteriophage K10 with its receptor, the lamB protein of Escherichia coli; Roa M; The lamB protein of Escherichia coli was initially recognized as the receptor for bacteriophage lambda . It is now shown also to constitute the receptor for phage K10 . The lamB protein interacts with phage K10 in vitro, but this interaction does not lead to phage inactivation . Most lambda-resistant labB mutants are also resistant to K10, and vice versa . However, a significant proportion of the mutants resistant to one of the phages is sensitive to the other . Nineteen K10-resistant lambda-sensitive mutants have been studied . Only six of them produce a lamB protein which seems totally unimpaired in its ihe same deletion interval of the lamB gene . The corresponding region of the lamB polypeptide must be specifically involved in the interaction with phage K10 . An unusual pattern of K10 host range mutants has been obtained; two calsses of such mutants could be defined, growing on two distinct classes of K10-resistant lamB mutants. J Bacteriol, 1979 Nov, 140(2), 351 - 8 Defective transport and other phenotypes of a periplasmic "leaky" mutant of Escherichia coli K-12; Anderson JJ et al.; A mutant of Escherichia coli K-12 deficient in high-affinity leucine transport and related binding proteins was obtained by selecting for azaleucine resistance after bacteriophage Mu mutagenesis . We determined that the cause was a generalized loss of periplasmic binding proteins and a sharp decrease in the activity of transport systems requiring them . Other transport systems resistant to osmotic shock and present in membrane vesicles, were affected to a lesser degree or not at all . The mutation, designated lky::Mucts, was shown to be a pleiotropic envelope mutation, rendering the mutant sensitive to ionic and nonionic detergents, antibiotics, and ethylenediaminetetraacetic acid: the strain had also acquired tolerance to colicins E1, E2, and E3, while remaining normally sensitive to a variety of bacteriophages . An analysis of the lipopolysaccharide of parent and mutant strains revealed a twofold reduction in the neutral sugar content of the core oligosaccharide of the lky strain, but no change in sensitivities to phages which utilize lipopolysaccharide or outer membrane proteins for absorption . The lky::Mucts locus was mapped by transduction and found to be located near, or in, the tolPAB gene cluster linked to gal . Secondary mutations suppressing the detergent sensitivity of lky arose at a frequency of 10(-7), yielding a variety of new phenotypes . The lky::Mucts mutation did not give rise to obvious alterations in the gross morphology of the cell or in cell division. J Virol, 1979 Nov, 32(2), 606 - 13 Electron microscopic analysis of partially replicated bacteriophage T7 DNA; Burck KB et al.; Partially replicated bacteriophage T7 DNA was isolated from Escherichia coli infected with UV-irradiated T7 bacteriophage and was analyzed by electron microscopy . The analysis determined the distribution of eye forms and forks in the partially replicated molecules . Eye forms and forks in unit length molecules were aligned with respect to the left end of the T7 genome, and segments were scored for replication in each molecule . The resulting histogram showed that only the left 25 to 30% of the molecules was replicated . Several different origins of DNA replication were used to initiate replication in the UV-irradiated experiments in which 32P-labeled progeny DNA from UV-irradiated phage was annealed with ordered restriction fragments of T7 DNA (K . B . Burck and R . C . Miller, Jr., Proc . Natl . Acad . Sci . U.S.A . 75:6144--6148, 1978) . Both analyses support partial-replica hypotheses (N . A . Barricelli and A . H . Doermann, Virology 13:460--476, 1961; Doermann et al., J . Cell . comp . Physiol . 45{Suppl.}:51--74, 1955) as an explanation for the distribution of marker rescue frequencies during cross-reactivation; i.e., replication proceeds in a bidirectional manner from an origin to a site of UV damage, and those regions of the genome which replicate most efficiently are rescued most efficiently by a coinfecting phage . In addition, photoreactivation studies support the hypothesis that thymine dimers are the major UV damage blocking cross-reactivation in the right end of the T7 genome. J Bacteriol, 1979 Nov, 140(2), 369 - 76 Cyclic adenosine 3',5'-monophosphate regulation of the bacteriophage T6/colicin K receptor in Escherichia coli; Alderman EM et al.; Mutant strains of Escherichia coli unable to synthesize cyclic adenosine 3',5'-monophosphate (cAMP) or the cyclic adenosine monophosphate receptor protein (CRP) were more resistant than wild-type cells to infection by bacteriophage T6 . This resistance was found to be associated with the decreased production of specific T6 receptor protein (also the colicin K receptor) located in the outer membrane protein fraction of these cells . Transcription of this particular outer membrane protein was regulated by the cAMP-CRP complex . A novel affinity technique coupled with sodium dodecyl sulfate-polyacrylamide gel electrophoresis was used in these investigations. Mol Gen Genet, 1979 Nov, 176(3), 369 - 74 Transcription and membrane attachment of bacteriophage lambda DNA in the absence of N function in the E . coli suA 1 mutant; Bourguignon-Van Horen MF et al.; In the polarity suppressor strain psuA 1, we observe a partial N independence of both transcription and DNA-membrane attachment for a lambda NN mutant . These results, in agreement with the genetical data reported by Dambly et al . (1976), suggest that the N product and rho factor are involved in the same process but may not interact directly. Gene, 1979 Nov, 7(3-4), 217 - 70 A comprehensive molecular map of bacteriophage lambda; Szybalski EH et al.; Physical and genetic mapping of deletion mutations has been correlated with the available molecular sizes of the lambda gene products and the DNA base sequence to construct a comprehensive molecular map of the phage lambda genome . The physical length of the DNA making up the left arm from the cos site through gene J is not sufficient to account in a nonoverlapping manner for all the proteins of the sizes reported to be coded, especially in the Nu1--C region . In the right arm all the coding capacity has not been accounted for, and it appears to be oversaturated only in the gam-ral region . The positions of several IS and Tn elements, and of restriction endonuclease cleavage sites are specified. J Virol, 1979 Nov, 32(2), 629 - 39 Electrophoretic characterization of intracellular forms of bacteriophage phi X174 DNA: identification of novel intermediate of altered superhelix density; Johnson PH et al.; The replication cycle of bacteriophage phi X174 DNA has been analyzed by agarose gel electrophoresis . The electrophoretic behavior of the predominant species of parental and progeny DNA molecules formed between 5 and40 min after infection was deduced and quantitated . Migration through 1.4% agarose at 5 and 10 V/cm resolved all known viral DNA species as well as fragments of host chromosomal DNA . Among parental replicative form(RF) molecules synthesized, 1 to 3% were full length linear duplexes (RFIII) and approximately 65% were closed circular duplexes (RFI) . Most of the input viral strands remained in a duplex structure throughout the period of infection studied here . Among progeny molecules, RFIII was not readily detected unless viral DNA synthesis was inhibited by chloramphenicol . Late in infection, 20% of the progeny RF were found to exist as form I dna . in addition, approximately 1% of the viral DNA was found as unit length linear single strands . Electrophoretic analysis of RF DNA after controlled denaturation suggests the existence of four populations of closed circular RF: (i) molecules of native superhelix density (RFI); (ii) a population of molecules of altered topological linking number, alpha, differing in increments of one superhelical turn (tau) between tau values of 0 and approximately -31; (iii) a superimposed population of topological isomers which under electrophoresis conditions have mean tau value (tau) equal to +5; and (iv) a population of "complexed" molecules with a reduced number of superhelical turns due to their association with single-stranded DNA and RNA . Complexed parental molecules isolated from cells infected at high multiplicity released FRI and homologous single-stranded DNA upon denaturation and are postulated to be intermediates in genetic recombination . Complexed RF DNA isolated from cells infected at low multiplicity release native supercoils upon reaction with RNase H and are observed by electron microscopy to contain displacement loops . Such molecules are likely intermediates in transcription . Our results are consistent with a structure of complexed RFI involving a partially triple-stranded helix in which a covalently closed circular duplex molecule contains a reduced number of superhelical turns due to the unwinding produced by base pairing between one strand of the supercoil and an associated homologous single strand of DNA or RNA. J Bacteriol, 1979 Nov, 140(2), 479 - 89 Lambda transducing bacteriophage carrying deletions of the argCBH-rpoBC region of the Escherichia coli chromosome; Linn T et al.; Deletions in the rpoBC region have been transferred to phage lambda and characterized in detail by genetic, structural, and functional tests . We thus extend and confirm knowledge of the organization of this part of the chromosome . The new phages are useful tools for studying the genes for the bacterial transcription and translation machinery. CRC Crit Rev Biochem, 1979 Nov, 7(1), 23 - 43 The enzymatic replication of DNA; Kornberg A; Enzymatic mechanisms of DNA replication have been investigated using small bacteriophages as probes to illuminate the cellular systems upon which they must rely during infection . Conversion of the circular, single-stranded DNAs of phages M13, G4, and phi X174 to their duplex forms has revealed the participation of diverse ways to start a new chain and a complex DNA polymerase III holoenzyme upon which all these systems depend for chain elongation . The phi X174 system, which is the most exacting and revealing of the host chromosomal replication pattern, includes at least twenty polypeptides for making the viral DNA into a duplex and multiplying the duplex . Resolution and purification of these numerous proteins is in train and their reconstitution into a "replisome"-like structure is envisioned. J Bacteriol, 1979 Nov, 140(2), 525 - 31 Defective F pili and other characteristics of Flac and Hfr Escherichia coli mutants resistant to bacteriophage R17; Burke JM et al.; Mutants resistant to the donor-specific bacteriophage R17 were isolated from Hfr and Flac-containing strains of Escherichia coli K-12 . Thirty-five mutants were examined for the presence of F pili by electron microscopy . The pilus morphology was studied, as were the abilities of the cells to retract their pili and to synthesize new pili . Measurements were made of the efficiency of the conjugal deoxyribonucleic acid transfer and of M13 and R17 phage infection . All mutants had noticeable defects in pilus production, structure, or function . Mutants were found which produced unusually long pili, displayed wide variations in the number of pili per cell, and were deficient in pilus retraction and synthesis . Evidence is presented that there may be two pathways of pilus retraction. Biochim Biophys Acta, 1979 Oct 25, 564(3), 495 - 506 Host-cell reactivation of alkylated T7 bacteriophage; Lane D et al.; Purified T7 phage, treated with methyl methanesulfonate, was assayed on Escherichia coli K-12 host cells deficient in base excision repair . Phage survival, measured immediately after alkylation or following incubation to induce depurination, was lowest on a mutant defective in the polymerase activity of DNA polymerase I (p3478) . Strains defective in endonuclease for apurinic sites (AB3027, BW2001) gave a significantly higher level of phage survival, as did the strain defective in the 5'--3' exonuclease activity of DNA polymerase I (RS5065) . Highest survival of alkylated T7 phage was observed on the two wild-type strains (AB1157, W3110) . These results show that alkylated T7 phage is subject to repair via the base excision repair pathway. J Biol Chem, 1979 Oct 25, 254(20), 10483 - 9 Characterization of the ribonucleic acid primers and the deoxyribonucleic acid product synthesized by the DNA polymerase and gene 4 protein of bacteriophage T7; Romano LJ et al.; The DNA polymerase and gene 4 protein of phage T7, in the presence of helix-destabilizing protein (DNA binding protein), catalyze DNA synthesis on duplex templates . As has been previously shown (Kolodner, R . D., and Richardson, C . C . (1978) 4 . Biol . Chem . 253, 574-584), in the absence of ribonucleoside 5'-triphosphates DNA synthesis is initiated at nicks, and all of the newly synthesized DNA is covalently attached to the template . In this paper we characterize the DNA synthesized in the presence of ribonucleoside 5'-triphophates and show that, in contrast, the major portion of the newly synthesized DNA is not attached to the template, having an average chain length of 5000 to 6000 nucleotides . In addition, each chain of newly synthesized DNA is terminated at its 5'-end by a covalently attached tetranucleotide RNA primer whose sequence is predominantly pppApCpCpC and pppApCpCpA . The results of isotope transfer experiments are in agreement with the number of initiation events determined by the incorporation of {gamma-32P}ATP and indicate that each of the four deoxyribonucleotides is present at the RNA-DNA junction. J Biol Chem, 1979 Oct 25, 254(20), 10476 - 82 Requirements for synthesis of ribonucleic acid primers during lagging strand synthesis by the DNA polymerase and gene 4 protein of bacteriophage T7; Romano LJ et al.; DNA polymerase and gene 4 protein of bacteriophage T7 catalyze DNA synthesis on duplex DNA templates . Synthesis is initiated at nicks in the DNA template, and this leading strand synthesis results in displacement of one of the parental strands . In the presence of ribonucleoside 5'-triphosphates the gene 4 protein catalyzes the synthesis of oligoribonucleotide primers on the displaced single strand, and their extension by T7 dna polymerase accounts for lagging strand synthesis . Since all the oligoribonucleotide primers bear adenosine 5'-triphosphate residues at their 5' termini, {gamma 32P}ATP is incorporated specifically into the product molecule, thus providing a rapid and sensitive assay for the synthesis of the RNA primers . Both primer synthesis and DNA synthesis are stimulated 3- to 5-fold by the presence of either Escherichia coli or T7 helix-destabilizing protein (DNA binding protein) . ATP and CTP together fully satisfy the requirement for rNTPs and provide maximum synthesis of primers and DNA . Provided that T7 DNA polymerase is present, RNA-primed DNA synthesis occurs on either duplex or single-stranded DNA templates and to equal extents on either strand of T7 DNA . No primer-directed DNA synthesis occurs on poly(dT) or poly(dG) templates, indicating that synthesis of primers is template-directed. Biochemistry, 1979 Oct 16, 18(21), 4581 - 8 In vitro transcription by wheat germ ribonucleic acid polymerase II: effects of heparin and role of template integrity; Dynan WS et al.; Double-stranded deoxyribonucleic acid (DNA) from bacteriophage lambda is a good template for wheat germ DNA-dependent ribonucleic acid (RNA) polymerase II . We delineated conditions for obtaining maximum polymerase activity using as template both the relatively intact DNA extracted from the the lambda phage and DNA into which single-strand nicks have been introduced by deoxyribonuclease (DNase) I digestion . The deliberate introduction of nicks produces a modest increase in transcription . The NaCl and MgCl2 optima are broader with the nicked template, so that higher concentrations of these salts are needed before polymerase activity begins to decline . Heparin inhibits initiation but not elongation by wheat germ polymerase . Polymerase can be protected against heparin inhibition by forming binary complexes with the template . The formation of these complexes is reduced at low temperature . The complexes, once formed, decay in the presence of heparin with a half-life of 10--20 min . The number of complexes is highly dependent on the degree of nicking of the template, suggesting that single-strand nicks are the predominant type of site where these heparin-resistant complexes are formed . Our data do not allow us to decide whether or not the presence of nicks plays as decisive a role in the absence of heparin. Eur J Biochem, 1979 Oct 15, 100(2), 433 - 40 Purification and some properties of deoxyribonuclease whose synthesis is controlled by gene 49 of bacteriophage T4; Nishimoto H et al.; An enzyme which specifically cleaves very-fast-sedimenting DNA of bacteriophage T4 is synthesized after infection of T4, and its synthesis is controlled by gene 49 {1,2} . This enzyme has been proved to be a DNase {2} . We have purified this DNase 3000-fold from extracts of E . coli infected with T4 . The purified preparation was practically free from other DNases, and the DNase activity was not detectable in cells infected with a mutant defective in gene 49 . The enzyme activity from cells infected with a temperature-sensitive mutant of gene 49 was also temperature-sensitive, suggesting strongly that gene 49 is a structural gene of the DNase . The molecular weight of the wild-type enzyme was estimated to be 50 x 10(3) by gel filtration chromatography . The purified DNase did not cleave native and denatured DNAs of T3 and T4, but cleaved renatured T3 DNA with enzymatically fragmented T3 DNA, indicating that gaps in the DNA duplex are structures susceptible to the DNase . Cleavage of the hybridized T3 DNA occurred when the fragmented DNA was phosphorylated at either the 3' or 5'-strand termini. Nature, 1979 Oct 11, 281(5731), 456 - 61 T4 DNA topoisomerase: a new ATP-dependent enzyme essential for initiation of T4 bacteriophage DNA replication; Liu LF et al.; A novel ATP-dependent DNA topoisomerase which makes reversible double-strand breaks in the DNA double helix has been purified to near homogeneity from T4 bacteriophage-infected Escherichia coli cells . Genetic data suggest that this activity is essential for initiating T4 DNA replication forks in vivo. J Biol Chem, 1979 Oct 10, 254(19), 9565 - 72 Purification of the T4 gene 32 protein free from detectable deoxyribonuclease activities; Bittner M et al.; Detailed procedures are presented which allow reproducible preparation of T4 gene 32 protein, a helix-destabilizing protein essential for DNA replication and genetic recombination in T4 bacteriophage-infected Escherichia coli cells . Although 32 protein can be purified to better than 99% homogeneity by any one of several procedures, these methods have been developed to remove trace amounts of contaminating deoxyribonucleases, which are present in high levels in the original infected cells . Two alternative preparations are presented, each involving three chromatographic steps . Both 32 proteins obtained are essentially "nuclease-free," when tested at physiological salt concentrations . However, we show here that the phenyl-Sepharose chromatography step, which is necessary to remove an exonuclease activity active only at low salt concentrations, also removes a second protein present in trace amounts . In some cases, retention of this second protein is desirable, since it is essential for obtaining RNA primed, de novo DNA chain starts in an in vitro DNA replication system, when this system is constructed by mixing highly purified preparations of each of the six replication proteins coded for by T4 genes 32, 43, 44, 62, 45, and 41. J Biol Chem, 1979 Oct 10, 254(19), 9416 - 28 Role of polymeric forms of the bacteriophage phi X174 coded gene A protein in phi XRFI DNA cleavage; Ikeda JE et al.; Gene A of the phi X174 genome codes for two proteins, A and A* (Linney, E.A., and Hayashi, M.N . (1973) Nature New Biol . 245, 6-8) of molecular weights 60,000 and 35,000, respectively . The phi X A* protein is formed from a natural internal initiator site within the A gene cistron while the phi X A protein is the product of the entire A gene . These two proteins have been purified to homogeneity as judged by sodium dodecyl sulfate polyacrylamide gel electrophoresis . Previous studies have shown that the phi X A protein is an endonuclease which specifically introduces a discontinuity in the A cistron of the viral strand of supertwisted phi XRFI DNA . In addition to this activity, the phi X A protein also causes relaxation of supertwisted phi XRFI DNA and formation of a phi XRFH DNA . phi X A protein complex which has a discontinuity in the A cistron of the viral strand . This isolatable complex supports DNA synthesis when supplemented with extracts of uninfected Escherichia coli which lack phi X A protein and phi XRFI DNA . The phi XRFII DNA . phi X A protein complex can be attacked by exonuclease III but is not susceptible to attack by E . coli DNA polymerase I, indicating that the 5'-end of the complex is blocked . Attempts to seal the RFII structure generated from the phi XRFII DNA . phi X A protein complex with T4 DNA ligase in the presence or absence of DNA polymerase were unsuccessful . The phi X A protein does not act catalytically in the cleavage of phi XRFI DNA . Under conditions leading to the quantitative cleavage of phi XRFI DNA, the molar ratio of phi XRFI DNA to added phi X A protein was approximately 1:10 . At this molar ratio, cross-linking experiments with dimethyl suberimidate yielded 10 distinct protein bands which were multiples of the monomeric phi X A protein . In the absence of DNA or in the presence of inactive DNA (phi XRFII DNA) no distinct protein bands above a trimer were detected . We found it possible in vitro to form a phi XRFII DNA . phi X A protein complex with wild-type phi XRFI DNA (phi X A gene+) and with phi XRFI DNA isolated from E . coli (su+) infected with phage phi X H90 (an am mutant in the phi X A gene) . Thus, in vitro, in contrast to in vivo studies, phi X A protein is not a cis acting protein . The purified phi X A* protein does not substitute for the phi X A protein in in vitro replication of phi XRFI DNA nor does it interfere with the action of the phi X A protein which binds only to supertwisted phi XRFI DNA . In contrast, the phi X A* protein binds to all duplex DNA preparations tested . This property prevents nucleases of E . coli from hydrolyzing duplex DNAs to small molecular weight products. Mol Gen Genet, 1979 Oct 3, 176(2), 221 - 31 Some properties of the chloramphenicol resistance transposon Tn9; Chandler M et al.; We have isolated variants of the plasmid RTF which have received the transposon Tn9 from bacteriophage P1Cm . We have shown by the formation of heteroduplex molecules between one RTF:Tn9 derivative and R100.1 that Tn9 is homologous to the r-determinant region of R100.1 which carries the determinants for chloramphenicol resistance . This suggests that Tn9 was derived from an r-det like structure by deletion, possibly mediated by one of the flanking IS1 elements . In spite of the similarity in structure between Tn9 and r-det however, we have demonstrated two distinct differences in the behavior of these two elements: 1) Tn9 but not r-det, is able to amplify, by a recA dependent mechanism, when cells harboring RTF::Tn9 are grown in the presence of chloramphenicol, and 2) Tn9, unlike r-det, does not form extrachromosomal circular molecules when RTF::Tn9 is tegrated into the bacterial chromosome. Mol Gen Genet, 1979 Oct 3, 176(2), 293 - 5 The origin of the DNA in transducing particles of bacteriophage Mu . Density gradient analyses of intact phages; Teifel J et al.; The origin of DNA in transducing particles of bacteriophage Mu was investigated by density labelling techniques . Unlabelled plaque-forming and leu+-transducing particles were of about the same density . Preinfection labelling of DNA with 5-bromodeoxyuridine increased the density of the transducing particles, but not that of the infective ones . Postinfection labelling increased the density of the infective particles twice as much as that of the transducing particles . We conclude that half of the transducing DNA is synthesized before infection and half is synthesized after infection, similar to the results obtained with P1kc transducing phages (Ikeda and Tomizawa, 1965). Mol Gen Genet, 1979 Oct 3, 176(2), 155 - 60 Influence of SOS repair on the specificity of radiation mutagenesis in bacteriophage phiX174; Bleichrodt JF et al.; The ochre mutant oc9 of bacteriophage phiX174 was irradiated with gamma-rays and the revertants were assayed on unirradiated and UV-irradiated host bacteria carrying an amber suppressor . The yield of revertants (amber + wild type) was higher on UV-irradiated than on unirradiated bacteria, showing that gamma-irradiated phiX174 was subjected to W-mutagenesis . For oc9 gamma-irradiated in the presence of oxygen the fraction of amber mutants among the revertants was lower when mutants were scored on UV-irradiated bacteria than when assayed on unirradiated indicator cells . The same fraction of ambers was obtained when mutants were assayed on unirradiated and UV-irradiated samples of a recA indicator strain . UV-irradiated phiX174 showed a similar phenomenon . These results suggest that the specificity with regard to insertion of bases opposite radiation damage in phiX174 DNA is different for host cells in which SOS repair has been induced and cells in which SOS repair is not operative. J Virol, 1979 Oct, 32(1), 268 - 75 Structural and biological properties of mycoplasmavirus MVL3: an unusual virus-procaryote interaction; Haberer K et al.; The kinetics of adsorption and growth of mycoplasmavirus MVL3 in Acholeplasma laidlawii 1305/68 host cells have been studied with one-step growth, premature lysis, and single-burst experiments . The virus was found to kill infected host cells . Virus release starts 90 min after infection and continues for about 10 to 15 h . Hence, virus production is unlike the classical lytic bacteriophages and instead resembles nonlytic cytocidal animal viruses . Structural details of the virus are described, and the molecular weight of the viral linear DNA has beenfound to be 26 x 10(6). Biochem J, 1979 Oct 1, 183(1), 65 - 71 Inhibition of bacteriophage-Q beta ribonucleic acid polymerase by guanosine triphosphate analogues; Brooks RR; Sixteen compounds related to GTP were evaluated as inhibitors of bacteriophage-Q beta poly(C)-dependent poly(G) polymerase . Non-phosphorylated compounds, including guanine, guanosine and deoxyguanosine, were inactive . Phosphorylated compounds gave significant inhibition at millimolar concentrations . For nucleotides the feature important for inhibition was the 5'-phosphate chain . Four triphosphates, XTP, ITP, 7-methyl-GTP and 2'-O-methyl-GTP, gave 50% inhibition of both the poly(C)- and poly(U2,C)-dependent reactions at concentrations from 0.1 to 5 mM . XTP was 10-fold more potent an inhibitor of the reaction with poly(U2,C) as template . None of these four compounds was able to substitute for GTP as substrate to a significant extent . The most active compound, 2'-O-methyl-GTP, was a competitive inhibitor (Ki = 0.4 mM) of GTP in the poly(C)-dependent reaction. Cell, 1979 Oct, 18(2), 247 - 56 Binding of mammalian ribosomes to MS2 phage RNA reveals an overlapping gene encoding a lysis function; Atkins JF et al.; The main binding site for mammalian ribosomes on the single-stranded RNA of bacteriophage MS2 is located nine tenths of the way through the coat protein gene . Translation initiated at an AUG triplet in the +1 frame yields a 75 amino acid polypeptide which terminates within the synthetase gene at a UAA codon, also in the +1 frame . Partial amino acid sequence analysis of the product synthesized in relatively large amounts by mammalian ribosomes confirms this assignment of the overlapping cistron . The same protein is made in an E . coli cell-free system, but only in very small amounts . Analysis of the translation products directed by RNA from op3, a UGA nonsense mutant of phage f2, identifies the overlapping cistron as a lysis gene . In this paper we show that the op3 mutation is a C yield U transition occurring in the second codon of the synthetase cistron, which explains the lowered production of phage replicase (as well as lack of lysis) upon op3 infection of nonpermissive cells . We discuss the properties of the overlapping gene in relation to its lysis function, recognition of the lysis initiator region by E . coli versus eucaryotic ribosomes and op3 as a ribosome binding site mutant for the f2 synthetase cistron. Cell, 1979 Oct, 18(2), 235 - 46 Characterization of Op3, a lysis-defective mutant of bacteriophage f2; Model P et al.; We have isolated a conditional lethal mutant of bacteriophage 12 which makes plaques only on E . coli strains carrying a UGA suppressor . It grows normally in nonsuppressing hosts but does not lyse such strains . The mutation complements with amber mutations in each of the three known phage cistrons . These observations lead us to postulate the existence of a fourth gene in the RNA phage. Ann Microbiol (Paris), 1979 Oct, 130B(3), 275 - 85 Restoration of DNA synthesis at non-permissive temperature and of UV resistance induced by bacteriophage Mu in Escherichia coli lig ts7 strain; Ghelardini P et al.; Infected by Mu, a mutant of E . coli, carrying the mutation lig ts7 shows a UV sensitivity intermediate between that of the uninfected lig-strain and that of the parental lig+ strain . At non-permissive temperature bacterial DNA replication is restored upon infection . These results suggest that DNA-ligase activity is coded for by the phage Mu genome. Biokhimiia, 1979 Oct, 44(10), 1864 - 76 {Conformational rearrangements of bacteriophage T4 lysozyme during its binding to the inhibitor}; Trontskii AV et al.; The structural changes of bacteriophage T4 lysozyme during its binding to the inhibitor, i . e . disaccharide-tetrapeptide N-acetylglucosaminyl-N-acetylmuraminyl - L - alanyl-gamma-D-glutaminyl - mesodiaminopimelyl-D-alanine) isolated from Escherichia coli cell wall have been studied . During the inhibitor binding to the protein the degree of helicity decreases by approximately 14% as was shown using the circular dichroism technique . The changes in optical properties of tryptophane, tyrosine and phenylalanine residues detected by UV difference and fluorescence spectroscopy have been observed . Based on the experimental data and a comparison of spatial organization of phage T4 lysozyme and chicken egg-white lysozyme made it possible to develop a structural model of phage T4 lysozyme functioning . This model may account for the differences in specificity of action of bacteriophage T4 and chicken egg-white lysozymes and allows to establish the role of the "extra" part of phage lysozyme . According to the model, at the first stage of binding the peptide part of the substrate comes in contact with the "upper" (with respect to the cleft) part of the protein molecule (residues 106--116 and 135--140) . This results in rearrangement of the molecule, with opening of the cleft at the second stage . This makes possible the access of the polysaccharide part of the substrate of the active site and a subsequent hydrolysis of the beta (1 leads to 4) glycoside bond. Proc Natl Acad Sci U S A, 1979 Oct, 76(10), 5110 - 3 Termination of transcription by bacteriophage T3 RNA polymerase: homogeneous 3'-terminal oligonucleotide sequence of in vitro T3 RNA polymerase transcripts; Majumder HK et al.; RNA was synthesized in vitro from a T3 DNA template by T3 RNA polymerase and subsequently separated into seven discrete size classes (molecular weights ranging between 0.21 x 10(6) and 6.2 x 10(6)) by electrophoresis in polyacrylamide slab gels . RNase T1-generated 3'-terminal oligonucleotide fragments were then selectively isolated from either the unfractionated total RNA or the gel-purified specific transcripts by chromatography on columns of dihydroxyboryl-cellulose . Sequence analysis of these oligonucleotide products indicated that the unfractionated transcripts as well as all the individual major RNA species examined had a unique sequence, (Gp)UpUpUpUpUpGOH, at their 3' termini . The specificity of this sequence, as well as the total lack of any sequence heterogeneity at the ends of these transcripts, indicates a high degree of specificity of termination during transcription in this system. Proc Natl Acad Sci U S A, 1979 Oct, 76(10), 4882 - 6 Transposition of the Escherichia coli insertion element gamma generates a five-base-pair repeat; Reed RR et al.; We have determined DNA sequences surrounding the termini of the Escherichia coli insertion element gamma delta, both at its normal locus on the F (fertility) factor and at three different sites of insertion into the plasmid pBR322 . After transposition, a five-base-pair pBR322 sequence is duplicated and appears in direct orientation adjacent to each end of the element . No such duplication flanks the ends of gamma delta in F, and there is no apparent homology between the sequences surrounding gamma delta in F and the five-base-pair duplications generated by insertion . These findings suggest that the duplications are not essential for transposition and that they do not act to direct gamma delta to a homologous site in the target chromosome . In addition, we find that the 35-base-pair inverted repeat that comprises the termini of gamma delta is strikingly similar in sequence to the ends of both the ampicillin-resistance transposon Tn3 and a 200-nucleotide-long sequence on the plasmid pSC101 which has been shown to mediate recombination with phage f1 replicative form . Within the terminal region, there is a specific heptanucleotide sequence common to each of the above elements and to bacteriophage Mu, all of which generate five-base-pair repeats upon insertion. Proc Natl Acad Sci U S A, 1979 Oct, 76(10), 4852 - 6 Gene 2 protein of bacteriophage T7: purification and requirement for packaging of T7 DNA in vitro; LeClerc JE et al.; The gene 2 protein of bacteriophage T7 is required for a late stage of T7 DNA replication because T7 gene 2 mutants fail to form normal concatemeric structures during the processing of newly synthesized T7 DNA . Extracts of gene 2 mutant phage-infected cells are unable to package T7 DNA into phage heads to form viable phage, as determined by an in vitro packaging assay for T7 DNA . Packaging activity can be stimulated greater than 100-fold in mutant extracts by the addition of extract prepared from cells infected with phage carrying a wild-type T7 gene 2, thus providing a complementation assay for the gene 2 protein . With this assay, the gene 2 protein has been purified to approximately 50% homogeneity . Purified preparations of the protein inhibit the activity of Escherichia coli RNA polymerase but have little effect on the activity of T7 RNA polymerase but have little effect on the activity of T7 RNA polymerase . The requirement for the gene 2 protein during T7 DNA replication may involve inactivation of E . coli RNA polymerase because the antibiotic rifampicin, a specific inhibitor of E . coli RNA polymerase, can substitute for the gene 2 protein in the in vitro packaging assay. J Virol, 1979 Oct, 32(1), 293 - 303 Electron microscopic mapping of RNA transcribed from the late region of polyoma virus DNA; Manor H et al.; The polyoma virus (Py) RNA species transcribed from the L DNA strand of the "late" region of the Py genome in Py-infected mouse cells have been mapped by hybridization with specific fragments of Py DNA followed by electron microscopic visualization of the hybrids . Total cellular polyadenylated Py-specific RNA molecules having an S value in the range of 16S to 20S were purified by oligodeoxythymidylic acidcellulose column chromatography, preparative hybridization with Py DNA, and sucrose gradient centrifugation . Cytoplasmic Py-specific RNA was similarily purified, except that it was not fractionated by sucrose gradient centrifugation . Hybrids of these RNA molecules and Py DNA fragments were spread for electron microscopy by either the cytochrome c technique or the bacteriophage T4 gene 32 protein method . The polyadenylic acid at the 3'-end of the RNA in the hybrids was identified by labeling with simian virus 40 DNA circles to which polybromodeoxyuridylic acid tails had been covalently attached . These experiments revealed the presence of three L DNA strand transcripts in both RNA preparations . Two of these RNA molecules were found to be spliced from chains transcribed from two noncontiguous parts of the late region . The third molecule either is a continuous transcript of the entire late region or contains a splicing feature which is too small to be reliably observed by the electron microscope methods used . The 5'-ends of the three RNA species map within a region extending from 68 to 70 map units on the Py restriction endonuclease map . Each of the two spliced molecules contains a 5'-terminal leader sequence transcribed from a DNA segment with an estimated length of 60 to 110 nuvleotides . The 3'-ends of the leaders map at 66.7 +/- 1.0 and 66.4 +/- 0.50 map units . In these molecules the 5'-ends of the other part (the main body) map at 59.4 +/- 0.90 and 49.4 +/- 2.0 map units, respectively . The 3'-termini of all three RNA species map at 24 to 25 map units. Genetics, 1979 Oct, 93(2), 285 - 96 The BAM H1 restriction site in the bacteriophage T4 chromosome is located in or near gene 8; Wilson GG et al.; Bacteriophage T4 cytosine-containing DNA is cleaved at a single site by the restriction endonuclease, Bam H1 . The site lies within the late region of the T4 genome, close to, or within, gene 8, one of the structural genes of the phage particle baseplate. J Bacteriol, 1979 Oct, 140(1), 83 - 91 A new gene of Escherichia coli K-12 whose product participates in T4 bacteriophage late gene expression: interaction of lit with the T4-induced polynucleotide 5'-kinase 3'-phosphatase; Cooley W et al.; We isolated five Escherichia coli mutants deficient in their ability to support the late (replication-coupled) gene expression of T4 bacteriophage at 30 degrees C . These mutants, which we call Lit mutants, define at least one novel gene at 25 min on the E . coli map . They were selected in an attempt to obtain mutants which restrict the growth of T4 mutants deficient in polynucleotide 5'-kinase 3'-phosphatase but not that of wild-type T4 at 37 degrees C . Some of the mutants do have these phenotypes under some conditions . Studies of the block in T4 development in some of the E . coli mutants suggest that Lit mutants are affected in a gene product involved in the metabolism of deoxyribonucleic acid nicks or single-strand gaps . None of the Lit mutants is deficient in the major, bacterial, 3'-phosphatase activity in crude extracts. J Virol, 1979 Oct, 32(1), 162 - 6 Heat-sensitive lambda repressors retain partial activity during bacteriophage induction; Lieb M; At 43 degrees C, lambda cIts prophages are "induced" and enter the lytic cycle . Lac- lysogens containing heat-inducible lambda N- prophages were superinfected with a lambda trp/lac N+cI- phage containing a lacZ+ gene whose expression is controlled by the lambda cI product (repressor) . Lysogens were then heated, and the synthesis of beta-galactosidase and release of progeny phage were measured . In lambda N- cIts2 or lambda N-cIts16 lysogens superinfected with lambda trp/lac N+, beta-galactosidase appeared earlier and was synthesized more rapidly than in superinfected lysogens containing lambda N- cIts857 prophage . Even at 45 degrees C, the cI857 repressor retained some activity . Lysogens containing other N-cIts mutant prophages producing renaturable repressors were also only partially derepressed at 43 degrees C . Partial derepression of lambda "early" transcription is sufficient for induction of lambda N+ prophages. Genetics, 1979 Oct, 93(2), 297 - 307 Some properties of site-specific and general recombination inferred from int-initiated exchanges by bacteriophage lambda; Echols H et al.; The site-specific recombination at the attachment site for prophage integration might proceed by two general mechanisms: (1) a concerted reaction without a free intermediate; (2) a sequential mechanism differing from typical general recombination only by an inability of the cross-strand intermediate structure to migrate into the region of nonhomology adjacent to the attachment site . The blocked-migration model predicts frequent genetic exchange in the int xis region near the attachment site if Int-mediated recombination occurs between lambda phage with homologous attachment sites . We find such additional int xis exchanges, but only at very low frequency (1% of the Int-mediated recombination) . We conclude that the resolution point only rarely moves away from the initial crossover point specified by Int and, therefore, that the Int reaction is mainly concerted . We interpret the rare additional int xis recombinants as indicative of occasional branch migration from an initial Int-mediated crossover . The frequency of the rare int xis recombinants is not simply related to distance from the attachment site to an int- or xis- mutation, suggesting that the heteroduplex distance is often at least a gene in length . The frequency of these additional exchanges is also not a strong function of distance between two mutations; from this we conclude that the resolution to the observed recombinant structure in the sequential cases occurs often by mismatch repair . We have found no marked effect of mutations in the bacterial recA, recB, recC, recF, or recL genes on the frequency of the int xis recombinants; this may indicate that none of these genes specifies a product uniquely required for resolution of a cross-strand intermediate. Mol Gen Genet, 1979 Oct 1, 175(3), 245 - 50 Loss of rac locus DNA in merozygotes of Escherichia coli K12; Evans R et al.; DNA-DNA hybridization was used to demonstrate that the substituted DNA in the bacteriophage lambda recE (formerly called lambda reverse) is homologous to DNA at the rac locus in Escherichia coli . Strains that are rac- do not contain appreciable amounts of this DNA, and it is lost from a rac+ episome (F' 123) after transmission to a rac- recipient . This is consistent with the proposal that the rac locus contains a cryptic prophage (Low, 1973). Proc Natl Acad Sci U S A, 1979 Oct, 76(10), 4877 - 81 Isolation and identification of bacteriophage phi X174 prohead; Mukai R et al.; The morphogenesis of bacteriophage phi X174 has been investigated by using an in vitro DNA synthesizing system . An extract of a B-mutant-infected cell is capable of synthesizing infectious phage in vitro when the B gene function is provided by the addition of an ammonium sulfate fraction of a C-mutant-infected-cell extract . This fraction contains the omega complex, a complex of phage-coded proteins with S = 108; the B-mutant extract does not . The purified omega complex, isolated from the C-mutant extract, caused the synthesis and encapsidation of viral DNA when added to B-mutant extract . The omega complex contains the B protein but it is the intact omega complex that functioned in the in vitro complementation of the B-mutant extract because other fractions containing B protein but no omega complex had little or no complementing activity . The results indicate that the omega complex is the phi X174 prohead . The B protein is not found in either the 132S or 114S phage particles but is an essential component of the prohead . This suggests that it may have a scaffolding function. J Bacteriol, 1979 Oct, 140(1), 294 - 6 Location of a ColE1 deoxyribonucleic acid region that affects the plaque-forming ability of lambda-ColE1 hybrid bacteriophage; Takeya T et al.; The plaque-forming ability of a hybrid phage between plasmidColE1 and phage lambda carrying amber mutations in genes O and P was inhibited by the presence of ColE1 in suppressor-deficient Escherichia coli cells . ColE1 deoxyribonucleic acid regions concerned with this inhibition were examined by using various deletion and transposon insertion derivatives of ColE1, and it was found that the presence of the deoxyribonucleic acid region extending between 420 and 613 base pairs upstream from the initiation site of ColE1 deoxyribonucleic acid replication (J . Tomizawa, H . Ohmori, and R . E . Bird, Proc . Natl . Acad . Sci . U . S . A . 74: 1865--1869, 1977) was essential for this function. J Gen Virol, 1979 Oct, 45(1), 195 - 200 Pilus-specific, lipid-containing bacteriophages PR4 and PR772: comparison of physical characteristics of genomes; Coetzee WF et al.; The genomes of pilus-specific, lipid-containing phages PR4 and PR772 were studied electron microscopically . An identical mol . wt . of 10.9 x 10(6) was obtained . The genomes are unique (non-permuted) and have cohesive ends . From the similarities in size and denaturation maps of the genomes and failue to demonstrate non-homology in heteroduplexes, reported morphological ambiguities were clarified . The known serological difference between the phages could not be related to non-homology of their genomes . It is concluded that phages PR4 and PR772 are the same phage. Mol Biol (Mosk), 1979 Sep-Oct, 13(5), 1064 - 9 {Kinetic parameters of superhelical DNA cleavage by endonuclease S1}; Gonikberg EM; Major kinetic parameters of endonuclease S1 were determined on superhelical bacteriophage PM2 DNA and on relaxed nicked circular PM2 DNA . At 37 degrees and 0,25 M NaCl, the Michaelis constants were respectively 1.7 . 10(-8) M and 1 . 10(-9) M, and catalytic constants were respectively 0.36 sec-1 and 1.2 . 10(-2) sec-1 . The inhibition of the enzyme reaction by its product was detected. J Invest Dermatol, 1979 Sep, 73(3), 236 - 8 Interstrand crosslinks in DNA of phage lambda after exposure to 8-methoxypsoralen and trimethylpsoralen in the presence of light; Cassuto E et al.; Two medically useful photosensitizing furocoumarins, 8-methoxypsoralen (8-MOP) and 4,5'8-trimethylpsoralen (TMP) were compared with respect to their abilities to produce interstrand crosslinks in DNA . DNA from bacteriophage lambda, labeled with 32P, was subjected to sedimentation in alkaline sucrose gradients following exposure to several concentrations of 1 of the 2 psoralens and irradiation (UV-A, 360 nm) for various times . In alkaline sucrose gradients, crosslinked DNA molecules sediment about 1.4 times faster than undamaged DNA strands and the proportion of molecules carrying crosslinks can be estimated with reasonable accuracy . At equimolar psoralen concentrations (2 x 10(-7)M, or 4 x 10(-5)M) and with increasing irradiation times, the rate of production of crosslinked DNA was 4 to 30 times greater for TMP than for 8-MOP . Differences in the therqpeutic efficacy of 8-MOP and TMP in various clinical situations may be accounted for by the types of photoadducts formed by each drug as well as by their solubilities, rates of absorption and rates of metabolic degradation. Gene, 1979 Sep, 7(1), 33 - 50 Thermo-inducible expression of cloned early genes of bacteriophage Mu; Giphart-Gassler M et al.; An EcoRI fragment, containing approx . 5100 base pairs (bp) of the immunity-end of bacteriophage Mu, was inserted into the multicopy plasmid pMB9 by in vitro recombination . The expression of early Mu genes, located on the cloned fragment, is thermo-inducible because of the presence of the ts mutation in gene c . The isolation of a transformant harbouring the recombinant plasmid, pGP1, was possible only when expression of Mu genes was prevented . pGP1 can be maintained at 28 degrees C at high copy number, but at 42 degrees C the pGP1 containing cells are killed due to the expression of the kil gene of Mu . The following Mu genes are present on pGP1: the ner gene, the integration and replication genes A and B, the cim gene, and the kil gene . pGP1 containing cells do not show Gam and Sot activity at 42 degrees C, therefore the leftmost EcoRI site on the Mu DNA is located between genes kil and gam or sot, or within the gam or sot gene. J Bacteriol, 1979 Sep, 139(3), 730 - 7 Two Escherichia coli chromosomal cistrons, sfrA and sfrB, which are needed for expression of F factor tra functions; Beutin L et al.; Twelve mutants of Escherichia coli K-12 have been isolated which carry chromosomal mutations that exhibit pleiotropic effects on the expression of F factor tra cistrons . F pilus synthesis, deoxyribonucleic acid transfer, and surface exclusion are all inhibited . Six of the mutants carry sfrA mutations, and six carry sfrB mutations . sfrA and sfrB are cistrons mapping near thr and metE, respectively . Several F-like plasmids are dependent on sfrA and on sfrB for expression of tra cistrons . Plasmids of incompatibility groups C and S are only dependent on sfrB,and other conjugative plasmids are dependent on neither . sfrB mutations also result in changes in certain cell envelope properties, including change sensitivity to certain bacteriophages which use lipopolysaccharide as a receptor, synthesis of nonfunctional flagella, and altered sensitivity to antibiotics. Proc Natl Acad Sci U S A, 1979 Sep, 76(9), 4294 - 8 Structure of genes for human growth hormone and chorionic somatomammotropin; Fiddes JC et al.; A 2.6-kilobase (kb) EcoRI restriction endonuclease fragment containing human growth hormone (hGH; somatotropin) gene sequences and a 2.8-kb EcoRI fragment containing human chorionic somatomammotropin (hCS; choriomammotropin) gene sequences have been identified by hybridization to cloned cDNA . Human DNA was cleaved with EcoRI and fractionated by preparative agarose gel electrophoresis; DNA in the size range 2--3 kb was ligated to lambda gt WES.lambda B DNA and viable recombinant bacteriophage were recovered by in vitro packaging . After infection of Escherichia coli and screening of phage plaques, single isolates of hGH and hCS gene sequences were obtained . Restriction endonuclease mapping showed that the hGH gene contains three intervening sequences interrupting the coding sequence . Partial DNA sequence analysis of the hGH gene, obtained by the chain termination method, confirmed the location of the intervening sequences and the identity of the fragment. Biophys J, 1979 Sep, 27(3), 447 - 53 Diffusion-controlled reactions on spherical surfaces . Application to bacteriophage tail fiber attachment; Bloomfield VA et al.; We have explored the kinetic implications of a model that may account for the acceleration of tail fiber (F) attachment to baseplates (B) by whiskers (W) on bacteriophage T4 . The model assumes that a W-F complex is formed initially, and that the tethered fiber then undergoes rotational diffusion until a B-F encounter takes place . In the absence of whiskers, B-F complexes must form unassisted . Formation of a W-F intermediate will accelerate F attachment to B if (a) the bimolecular rate constant for W-F complex formation is larger than that for direct B-F interaction and (b) subsequent rotational diffusion of the tip of F to B is not much slower than the dissociation of W-F . Condition a was investigated by applying a recent theory of orientational effects on translational diffusion-controlled reactions . This theory suggests that substantial rate enhancement is expected if the reaction half-angle theta 0 is larger for W-F than for B-F complex formation . Condition b was investigated by calculating the mean and the variance of the time required for the diffusion of a molecule (the proximal tip of the fiber) on a spherical surface (whose radius is the distance from the tip to the whisker tethering point) into a circular sink (the baseplate site) . The mean time is on the order of the inverse rotational diffusion coefficient, DR, of the fiber, but is sensitive to theta 0 . Both conditions are satisfied for plausible choices of parameters . The solution to the diffusion equation we have obtained should have application to other physical situations, such as the rate of quenching of a fluorophore as it diffuses on the surface of a spherical membrane into proximity with a quencher. Antimicrob Agents Chemother, 1979 Sep, 16(3), 421 - 3 Virucidal activity of retinal; Reinhardt A et al.; Herpes simplex virus type 2 and simian virus 40 were rapidly inactivated by retinal at micromolar concentrations . Other fat-soluble vitamins, particularly vitamin A derivatives, were also active against herpes simplex virus type 2 and several lipid-containing bacteriophages. Z Naturforsch {C}, 1979 Sep-Oct, 34(9-10), 811 - 4 T2 phage sensitization by linear and angular furocoumarins; Baccichetti F et al.; T2 bacteriophage sensitization has been studied using two furocoumarins capable of linking covalently to DNA to the same extent but producing different damages, psoralen and 4,5'-dimethylangelicin . Psoralen is a well-known linear furocoumarin capable of inducing in DNA both monoadducts and cross-links; 4,5'-dimethylangelicin is a new angular compound known as a pure monofunctional reagent . In the sensitization of T2 mature virions both drugs proved very active, yielding survival curves practically superimposable; on the contrary, in the experiments with the T2 vegetative form, i . e . its DNA inside the host, 4,5'-dimethylangelicin resulted much less effective, resembling the picture observed in the inactivation of the host bacteria . This result did not appear related to an enhancement of DNA repair by a Weigle effect . The different killing activity of 4,5'-dimethylangelicin can be explained supposing that this drug is capable of inducing cross-links in T2 DNA inside the virus core, in which it exists in a very folded form, but not in the same DNA after injection into the host bacteria. Proc Natl Acad Sci U S A, 1979 Sep, 76(9), 4530 - 3 Lactose genes fused to exogenous promoters in one step using a Mu-lac bacteriophage: in vivo probe for transcriptional control sequences; Casadaban MJ et al.; The lactose structural genes, without the lactose promoter, have been incorporated into the bacteriophage Mu genome to form a Mu-lac specialized transducing phage . This phage also carries a gene encoding resistance to ampicillin (Ap){Mu(Ap, lac)} . After infection and upon establishment of lysogeny, the Mu(Ap, lac) genome can integrate into apparently random sites in the Escherichia coli chromosome . When integration occurs within a gene in the orientation of its transcription, the lactose structural genes are so situated that they become expressed solely from the promoter of that gene . Thus, expression of the lactose genes of Mu(Ap, lac) can be used as an assay for transcription of that gene and for functional and mutational studies of gene regulation. Proc Natl Acad Sci U S A, 1979 Sep, 76(9), 4464 - 8 Cloning of integrated Moloney sarcoma proviral DNA sequences in bacteriophage lambda; Vande Woude GF et al.; We have identified integrated proviral DNA sequences of m1 and HT-1 isolates of Moloney sarcoma virus (MuSV) in EcoRI digests of transformed mink cell genomic DNA and have cloned these fragments in bacteriophage lambda . Both the lambda-HT1 phage recombinant, containing a 12.3-kilobase MuSV pair (kb) fragment, and the lambda-m1 phage recombinant, containing a 7.0-kb fragment, possess full copies of the sarcoma viruses along with 5' and 3' host flanking sequences . The MuSV proviral DNA sequences, 6.7 kb for HT-1 and 5.2 kb for m1, are colinear by heteroduplex microscopy with the 1.5-kb difference in size accounted for by two approximately equal to 0.8-kb deleted regions in m1 . Both integrated viral genomes are terminally redundant and have integrated at the same site in the provirus but at different sites on the host chromosome . The host sequence flanking integrated HT-1 MuSV have been identified as a single EcoRI restriction fragment of 5.6 kb in normal mink cells. Int J Radiat Biol Relat Stud Phys Chem Med, 1979 Sep, 36(3), 241 - 7 Alkali-labile sites and post-irradiation effects in gamma-irradiated biologically active double-stranded DNA in aqueous solution; Lafleur MV et al.; Gamma-irradiation of double-stranded RF-DNA of bacteriophage phi X174 in aqueous solution in the presence of oxygen produces at least one type of alkali-labile site . It is lethal and gives rise to breaks by alkali and is identical with the damage which becomes manifest by post-irradiation heat treatment . The effect of alkali is dependent on temperature . Furthermore, the excision repair system is not involved in eliminating lethal nucleotide damage in RFI-DNA. J Bacteriol, 1979 Sep, 139(3), 738 - 47 Lambda bacteriophage-mediated transduction of ColE1 deoxyribonucleic acid having a lambda bacteriophage-cohesive end site: selection of packageable-length deoxyribonucleic acid; Umene K et al.; An in vitro recombinant ColE1-cos lambda deoxyribonucleic acid (DNA) molecule, pKY96, has 70% of the length of lambda phage DNA . The process of lambda phage-mediated transduction of pKY96 generated a small amount of transducing phage particles containing ColE1-cos lambda DNA molecules of 80 or 101% of the length of lambda phage DNA, in addition to those containing original pKY96 DNA molecules . The newly isolated larger plasmid DNAs were transduced 100 times more efficiently than pKY96 DNA . Their structures were compared with that of a prototype pKY96 DNA, and the mechanism of the formation of these molecules is discussed. J Biol Chem, 1979 Aug 25, 254(16), 7534 - 9 The properties of a bacteriophage T5 mutant unable to induce deoxyuridine 5'-triphosphate nucleotidohydrolase . Synthesis of uracil-containing T5 deoxyribonucleic acid; Warner HR et al.; Bacteriophage T5 induces a deoxyuridine 5'-triphosphate nucleotidohydrolase (dUTPase) activity during infection of Escherichia coli . A T5 mutant (T5 dut) unable to induce this dUTPase activity has been isolated . Although this mutant is viable, the E . coli dUTPase activity is not sufficiently active to exclude uracil from the progeny DNA and about 3% of the thymine is replaced by uracil . When the mutant is grown in an E . coli dut host about 12% of the thymine in the progeny DNA is replaced by uracil . T5 phage containing 12% uracil can replicate in uracil-DNA glycosylase-deficient (ung) hosts with high efficiency, but fail to replicate in ung+ hosts . The amount of thymine replaced by uracil in the progeny produced in dut hosts is nearly independent of the ung genotype, indicating that the host uracil-DNA glycosylase-dependent repair pathway is not operating efficiently to remove uracil from T5 progeny DNA. J Biol Chem, 1979 Aug 25, 254(16), 8042 - 51 The purification and properties of a double-stranded DNA-binding protein encoded by the gene D5 of bacteriophage T5; Rice AC et al.; We have purified a DNA-binding protein from bacteriophage T5-infected cells . The protein is the product of the T5 gene D5 and is produced in quantities ultimately exceeding 2% of the total cell protein . The protein has no te