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| J Bacteriol, 1976 Oct, 128(1), 56 - 68 Thermal adaptation in yeast: growth temperatures, membrane lipid, and cytochrome composition of psychrophilic, mesophilic, and thermophilic yeasts; Arthur H et al.; The temperature limits of growth of a number of yeast species were examined, and on this basis the organisms were classified into different thermal categories . The following species were examined: Leucosporidium frigidum and Leucosporidium nivalis, psychrophilic, temperature limits of growth, -2 to 20 degrees C; Canadian lipolytica mesophilic, temperature limits of growth, 5 to 35 degrees Candida parapsilosis and Saccharomyces telluris, thermotolerant, temperature limits of growth, 8 to 42 degrees C; Torulopsis bovina and Candida slooffi, thermophilic, temperature limits of growth, 25 to 45 degrees C and 28 to 45 degrees C, respectively . The membrane lipid and cytochrome composition of mitochrondrial fractions isolated from these yeasts were compared . There was a direct correlation between the growth temperature and the degree of membrane of lipid unsaturation; the lower the temperature, the greater the degree of lipid unsaturation . The membrane lipid composition of the thermophilic yeasts were distinguished by the high percentage (30 to 40%) of saturated fatty acid, as compared with the mesophilic and psychrophilic yeasts . The latter contained approximately 90% unsaturated fatty acid, 55% of which was linolenic acid, C alpha-18:3 . Changes in phospholipid composition in relation to temperature were also noted . The respiratory-deficient thermophile, C . slooffi, was characterized by the absence of cardiolipin (sensitivity 0.1 mug of phosphorus) and cytochrome aa3 . The absence of conventional mitochondrial structures in this thermophilic microorganism is tentatively suggested although low concentrations of cytochromes b, c, and c1 were detected by low-temperature spectroscopy . On the other hand, the respiratory-competent thermophile, T . bovina, was characterized by a high cardiolipin (25% of the total phospholipid) and cytochrome aa3 content (1 nmol/mg of mitochrondrial protein) . Low-temperature spectra showed the presence of one b-type cytochrome in the thermophilic yeasts, two b-type cytochromes in the mesophilic yeasts, and three b-type cytochromes in the psychrophilic yeasts . It was concluded that a knowledge of the properties of the biological membrane is fundamental to an understanding of the ability of a microorganism to grow and reproduce in different temperature environments. Biochim Biophys Acta, 1976 Sep 14, 445(2), 386 - 97 Purification and properties of extracellular alpha-glucosidase of a thermophile, Bacillus thermoglucosidus KP 1006; Suzuki Y et al.; An extracecular alpha-glucosidase (alpha-D-glucoside glycohydrolase, EC 3.2.1.20) of a thermophile, Bacillus thermoglucosidius KP 1006, was purified about 350-fold . The purified enzyme had a specific activity of 164 mumol of p-nitrophenyl-alpha-D-glucopyranoside hydrolyzed per min at 60 degrees C and pH 6.8 per mg of protein . The molecular weight was estimated at 55 000 . The pH and temperature optima for activity were 5.0--6.0 and 75 degrees C, respectively . Below 40 degrees C, the activity was less than 4.5% of the optimym . The enzyme showed a high specificity for alpha-D-glucopyranoside . The maximal hydrolyzing velocity per substrate diminished in the order: phenyl-alpha-D-glucopyranoside, p-nitrophenyl-alpha-D-glucopyranoside, isomaltose, methyl-alpha-glycopyranoside . The respective Km values were 3.0, 0.23, 3.2 and 27 mM . The activity was trace for turanose, and not detectable for sucrose, trehalose, raffinose, melezitose, maltose, maltotriose, phenyl-alpha-D-maltoside, dextran, dextrin and starch . Tris, p-nitrophenyl-alpha-D-xylopyranoside, glucose and glucono-delta-lactone blocked competitively the enzyme with respect to p-nitrophenyl-alpha-D-glucopyranoside . The Ki values were 0.12, 0.14, 2.2 and 2.4 mM, respectively . The activity was affected by heavy metal ions, but insensitive to EDTA, p-chloromercuribenzoate and iodoacetate . The enzyme was stable up to 60 degrees C, and inactivated rapidly at temperatures beyond 72 degrees C . The pH range for stability was 4.0--11.0 at 31 degrees C, and 6.0--8.5 at 55.5 degrees C . At 25 degrees C, the enzyme failed to be inactivated in 45% ethanol, in 7.2 M urea, and in 0.06% sodium dodecyl sulfate, but the tolerance was extremely reduced at 60 degrees C. Biochemistry, 1976 Sep 7, 15(18), 4026 - 33 Isolation and properties of 6-phosphogluconate dehydrogenase from Escherichia coli . Some comparisons with the thermophilic enzyme from Bacillus stearothermophilus; Veronese FM et al.; 6-Phosphogluconate dehydrogenase (6-phospho-D-gluconate:NADP oxidoreductase (decarboxylating), EC 1.1.1.44) of Escherichia coli MREp 600 has been isolated with the purpose of carrying out comparative studies with the thermostable enzyme previously isolated from Bacillus stearothermophilus (Veronese, F.M., Boccu, E.,Fontana, A., Benassi,C.A., and Scoffone, E . (1974), Biochim, Biophys . Acta 334, 31) . The purified enzyme appeared homogeneous by the criteria of disc gel electrophoresis with and without sodium dodecyl sulfate, ultracentrifugation, and gel filtration . The enzyme has enzymological and physiochemical properties similar to the enzyme isolated from other sources, including B . stearothermophilus . The E . coli enzyme has a mol wt of 100,000 +/- 3000 and is composed of two apparently identical subunits . The amino acid composition of both the mesophilic and thermophilic enzyme has been determined and found to present large similarities . The E . coli enzyme shows a high degree of specificity for nicotinamide adenine dinucleotide (NADP) and it is inhibited by reduced NADP (NADPH) . Cysteine residues are involved in the catalytic activity, since on incubation of the enzyme with p-chloromercuribenzoate or 5.5'-dithiobis(2-nitrobenzoic acid) strong inhibition occurs, activity being restored by treatment with excess of beta-mercaptoethanol . The substrate 6-phosphogluconate protects partially the enzyme from inactivation . Both the mesophilic and thermophilic 6-phosphogluconate dehydrogenases are inactivated by Rose Bengal in the presence of light by similar kinetics and protected against photoinactivation by the enzyme substrate . The E . coli enzyme, on the other hand, showed distinct differences in stability against heat and unfolding agents in respect to the B . stearothermophilus enzyme . Heating at 50 degrees C or incubation in 8 M urea results in rapid inactivation . The gross structure of the mesophilic and thermophilic enzyme was very similar as judged by circular dichroic measurements . The far-ultraviolet circular dichroic spectrum had a negative band centered at about 220 nm . In both cases, the fluorescence emission spectrum indicates that the environment of the tryptophan residues is similar, since both enzymes show an emission maximum at 334 nm upon excitation at 295 nm . Circular dichroism measured at various temperatures between 25 and 80 degrees C showed the mesophilic enzyme to be conformationally stable below about 45 degrees C and the thermophilic enzyme below 60 degrees C . The secondary structure of the E . coli enzyme was very sensitive to the denaturing action of urea, since in 8 M urea it rapidly unfolded . Partial renaturation after urea treatment occurred on dilution with buffer or dialysis, as evidenced by spectral properties of the renatured enzyme . The results show that the mesophilic and thermophilic enzymes are very similar and that differences in thermal stability depend on subtle differences in the architectures of the proteins. Postgrad Med, 1976 Sep, 60(9), 121 - 7 Allergic alveolar diseases . Problems in diagnosis and management; Richerson HB; One of the diagnostic possibilities to consider when a patient presents with cough, fever, dyspnea, or pulmonary infiltrates is hypersensitivity pneumonitis . Some of the problems encountered in diagnosis of diffuse lung disease are illustrated in two case reports . In one of the cases, interstitial pneumonitis of insidious onset was attributed to inhalation of thermophilic organisms in moldy silage . In the other, the outstanding pathologic feature was bronchiolitis obliterans, and circumstantial evidence pointed to a home humidifier as the source of the problem. J Biochem (Tokyo), 1976 Sep, 80(3), 485 - 90 Some catalytic and molecular properties of threonine deaminase from Bacillus stearothermophilus; Muramatsu N et al.; Threonine deaminase {EC 4.2.1.16} was highly purified from Bacillus stearothermophilus . The enzyme exhibited maximum activity at 65 degrees and at pH 9.2--9.6 . It was inactivated on dilution and on storage at 4 degrees, but was protected by egg albumin . The enzyme was labile at 65 degrees, but became stable in the presence of egg albumin and isoleucine at pH 7.0 . The substrate saturation curve for the enzyme reaction at 40 or 65 degrees was hyperbolic, but in the presence of isoleucine, the curve became sigmoidal (n = 2) . The enzyme was more sensitive to isoleucine at 40 degrees than at 65 degrees, while valine slightly inhibited the enzyme at both 40 and 65 degrees . Inhibition of the enzyme by isoleucine was antagonized by valine at 40 and 65 degrees . These properties were essentially similar to those of the enzymes from mesophilic and thermophilic bacteria . The enzyme existed in two forms with different molecular sizes, 1.5-5 X 10(6) and 2 X 10(5) daltons, at pH 7.0 and at temperatures below 40 degrees . The larger component disaggregated into the small one at pH 8.5 or above, at temperatures above 50 degrees or in the presence of isoleucine and valine. J Bacteriol, 1976 Sep, 127(3), 1550 - 7 Deoxyribonucleic acid polymerase from the extreme thermophile Thermus aquaticus; Chien A et al.; A stable deoxyribonucleic acid (DNA) polymerase (EC 2.7.7.7) with a temperature optimum of 80 degrees C has been purified from the extreme thermophile Thermus aquaticus . The enzyme is free from phosphomonoesterase, phosphodiesterase and single-stranded exonuclease activities . Maximal activity of the enzyme requires all four deoxyribonucleotides and activated calf thymus DNA . An absolute requirement for divalent cation cofactor was satisfied by Mg2+ or to a lesser extent by Mn2+ . Monovalent cations at concentrations as high as 0.1 M did not show a significant inhibitory effect . The pH optimum was 8.0 in tris(hydroxymethyl)aminomethane-hydrochloride buffer . The molecular weight of the enzyme was estimated by sucrose gradient centrifugation and gel filtrations on Sephadex G-100 to be approximately 63,000 to 68,000 . The elevated temperature requirement, small size, and lack of nuclease activity distinguish this polymerase from the DNA polymerase of Escherichia coli. J Bacteriol, 1976 Sep, 127(3), 1136 - 40 Synthesis of omega-alicyclic fatty acids from cyclic precursors in Bacillus subtilis; Dreher R et al.; A mutant of Bacillus subtilis synthesizes a variety of omega-alicyclic fatty acids when fed with the respective alicyclic carboxylic acids . These fatty acids are: omega-cyclopropane, omega-cyclobutane, omega-cyclopentane, omega-cyclohexane, and omega-cyclohexene fatty acids . These unusual fatty acids did not lead to an inhibition of growth at 37 degrees C and pH 7 . The selective advantage of these fatty acids under extrene conditions was studied in comparison with the acidophilic, thermophilic bacterium B . acidocaldarius, which normally contains a high proportion of omega-cyclohexane fatty acids. Mycopathologia, 1976 Aug 30, 59(1), 29 - 35 A study of fungal spore poupulations in the atmosphere of Kuwait; Moustafa AF et al.; The fungal air-spora of Kuwait was investigated using the Petri-dish trapping technique . During the period from 1st April, 1974 to 30th June, 1975, a total of 3685 colonies were recorded from 2% malt agar plates . Fifty-five genera and 116 species were identifed . Alternaria occupied the first place in the order of percentage incidence . being represented by 18.3% of the entire catch, followed by Aspergillus (17.1%), Penicillium (14%), Cladosporium (13.6%), Drechslera (13.3%), and Ulocladium (7.1%) . The widest spectrum of species was displaced by Aspergillus (possessed 19 species) followed by Penicillium (17 species), Alternaria (6 species), Drechslera (5 species), and Ulocladium (4 species) . The monthly total number of fungi showed marked seasonal periodicity with the greatest number of colonies occurring in March-April and November . The lowest counts were recorded in mid-summer (July) and mid-winter (January) . Only 220 colonies and 17 species of thermophilous fungi were reported from plates incubated at 45 degrees C . The most common species were Aspergillus fumigatus and A . terreus. Can J Microbiol, 1976 Aug, 22(8), 1102 - 12 The fine structure of conidial development in the genus Torula . IV . T . thermophila Cooney & Emerson; Ellis DH et al.; Torula thermophila produced typical chlamydospores either as intercalary chains within prostrate hyphae or as terminal swellings on short, lateral, hyphal branches . Mature chlamydo-spores were spherical, dark brown, smooth-surfaced structured structures with thick, single-layed cell walls (= secondary wall layer) usually differentiated into an outer electron-dense zone and an inner electron-transparent zone . Disarticulation and sport release occurred after the disintegration of the original hyphal wall . The thallospores of T . thermophila arise in a manner different from the blastospores produced by other species of Torula and are structurally more closely related to the spores produced by Humicola insolens . However until further work has been completed on spore development in the Torula-Humicola complex of fungi the name T . thermophila is retained. Biochim Biophys Acta, 1976 Jul 2, 435(3), 228 - 35 Purification and properties of the RNA polymerase of an extremely thermophilic bacterium: Thermus aquaticus T2; Fabry M et al.; The RNA polymerase of the extremely thermophilic bacterium Thermus aquaticus T 2 has been purified to homogeneity . The apparent molecular weights of the subunits of the enzyme are : 165 000, 130000, 92 000 and 44 000 . The in vitro temperature optimum of enzyme activity is around 65 degrees C . The enzyme has a preference towards the homologous template and is strongly inhibited by KCI . Rifampicin inhibits the enzyme only to 50% even at very high concentrations . Heparin inhibits it completely, but only at higher molar excess than in the case of the Escherichia coli enzyme . The enzyme can form heparin-resistant complexes at elevated temperatures on the homologous template, but not on E . coli DNA. Prikl Biokhim Mikrobiol, 1976 Jul-Aug, 12(4), 505 - 8 {Thermogenesis of mesophilic, thermotolerant and thermophilic strains of microorganisms--producers of fungal cell wall--destroying enzymes}; Koriagin VV et al.; Investigations were carried out to clarify the relationship between thermogenesis and production of yeast wall lyzing enzymes by the mesophilic strain of Bacillus subtilis, thermotolerant strain of Actinomyces sp . II and thermophilic strain of Actinomyces sp . 10 . The enzymic lyzing activity was measured in the culture liquid filtrate of those microorganisms . The thermophilic strain of Actinomyces sp . 10 showed the highest enzymic activity . The thermogenetic curves of the cultures had several inflections . The mesophilic culture of Bacillus subtilis whose enzymic lyzing activity was the lowest displayed the highest heat release. Appl Environ Microbiol, 1976 Jul, 32(1), 131 - 7 Study of plating efficiency of bacteriophages of thermophilic lactic acid bacteria on different media; Sozzi T et al.; The qualitative and quantitative abilities of phages of 13 strains of thermophilic lactic bacteria (lactobacilli and streptococci) to produce plaques on six different media were studied . The influence of the addition of calcium on the one hand, and of incubation conditions (aero-anaerobiosis), on the other hand, was also examined . For the phages of lactobacilli, the best results were obtained with the medium of de Man, Rogosa, and Sharpe (1960), whereas for the phages of streptococci the medium of Hogg and Jago (1970) appeared to be the best . Calcium and incubation conditions play a role which is variable in importance, but rarely negligible. Nucleic Acids Res, 1976 Jul, 3(7), 1703 - 13 CD spectra of 5-methyl-2-thiouridine in tRNA-Met-f from an extreme thermophile; Watanabe K et al.; 5-Methyl-2-thiouridine (S) in tRNA-Met-f from an extreme thermophile is located in the TpsiC region, replacing T, and has a positive CD band centered at 310 nm . Upon heating, the profiles of the change in this band were similar to the UV melting profiles of the change monitored at 260 nm . This strongly suggests a close relation between heat denaturation of the tRNA and the conformation of the S base . Oligonucleotides containing S showed negative CD bands at 320-330 nm, like the monomer S itself, but when the 3'-2/5 fragment containing S formed a complex with the complementary 5'-3/5 fragment, a positive CD band appeared at 310 nm . These results suggest that combination of the TpsiC loop containing S with the hU loop is necessary for the positive band of S at 310 nm . S may serve to strengthen the association of the TpsiC loop with the hU loop in tRNA of the thermophile. Am Rev Respir Dis, 1976 Jul, 114(1), 29 - 43 Cellular and humoral bronchopulmonary immune response of rabbits immunized with thermophilic actinomyces antigen; Harris JO et al.; The immunologic events involved in the pathogenesis of human hypersensitivity pneumonitis are poorly understood . To delineate better the cellular and humoral bronchopulmonary response to host challenge with an important agent for hypersensitivity pneumonitis, an animal model was developed . Rabbits intratracheally inoculated with Micropolyspora faeni antigen developed histologic lesions resembling hypersensitivity pneumonitis in man, characterized by a mononuclear cell interstitial reaction and a marked increase in the number of intra-alveolar cells . This reaction was intense 8 days after immunization and progressively decreased in intensity, with complete resolution occurring, in some instances, by 3 weeks . The increased intra-alveolar cells were predominantly macrophages, but soon after immunization, greater numbers of lymphocytes and granulocytes were present . Associated with the intra-alveolar macrophage response was the appearance of "activated" macrophages, based on increased glucose oxidation and ultrastructural appearance . The immunized rabbits demonstrated Arthus-type skin reactions, as well as delayed hypersensitivity skin reactions after intradermal injection of M . faeni antigen . Alveolar macrophage migration was significantly inhibited in immunized rabbits in the presence of M . faeni antigen, suggesting that specifically sensitized lymphocytes were present in the free bronchoalveolar cell population . Intratracheal inoculation with M . faeni resulted in the production of anti-M . faeni-precipitating antibodies in the sera of all animals and in bronchoalveolar secretions in 15 of 16 inoculated rabbits . The concentrations of IgG and IgA in lung-wash fluid were significantly greater in immunized animals . In the immunized rabbits, the mononuclear cell response, positive M . faeni-induced direct migration inhibition factor assays, and delayed skin reactivity strongly suggested the presence of a delayed hypersensitivity component in disease pathogenesis . The presence of precipitating antibody and the increased concentrations of immunoglobulins were consistent with stimulation of the humoral immune system; however, the role of humoral mechanisms in the pathogenesis of the pulmonary lesions is less certain than that of delayed hypersensitivity. Am Rev Respir Dis, 1976 Jul, 114(1), 23 - 8 The frequency of precipitins to trichloroacetic acid-extractable antigens from thermophilic actinomycetes in farmer's lung patients and asymptomatic farmers; Roberts RC et al.; The hypothesis that the trichloroacetic acid (TCA)-extractable antigen fractions prepared from Micropolyspora faeni cells and other thermophilic actinomycetes are better diagnostic immunodiffusion reagents for the detection of farmer's lung disease than the crude extracellular antigen concentrates has been tested . The frequency of serum precipitins to the TCA-extractable antigens has been compared in symptomatic farmer's lung patients and in healthy farmers having positive precipitin tests to the crude antigen concentrates of the analogous thermophilic actinomycetes . Sixty-eight per cent of the symptomatic group were found to have precipitins to the M . faeni TCA-extractable antigens compared with only 22 per cent of the asymptomatic group . With regard to the other thermophilic actinomycetes tested, no significant differences in frequencies of precipitins to TCA-extractable antigens between the symptomatic and asymptomatic groups were found . However, the total number of patients available for testing with each of these antigens was small . Sufficient chemical characterization of the TCA-extractable antigens was carried out to show that they were chemically similar to the TCA-extractable antigens prepared by other workers. J Biochem (Tokyo), 1976 Jul, 80(1), 141 - 51 Proton translocating ATPase of a thermophilic bacterium . Morphology, subunits, and chemical composition; Kagawa Y et al.; 1 . A stable membrane-bound ATPase {EC 3.6.1.3} (TF0-F1) capable of proton translocation in reconstituted vesicles was purified from the thermophilic bacterium PS3 cultured in medium containing L-{U-14C}amino acids . 2 . TF0-F1 was composed of a catalytic moiety (TF1) and a hydrophobic moiety (TF0) . TF1 contained 3 polypeptide chains with molecular weights of 56,000, 3 of 53,000, 1 of 32,000, 1 of 15,500, and 1 of 11,000 . TF0 contained 1 chain of 19,000, 2 of 13,500, and 5 of 5,400 daltons . TF1 was dissociated into subunits much less readily than F1 . 3 . TF1 consisted of 95A particles arrayed in hexagonal microcrystals . TF0-F1 consisted of a sphere (TF1) and a stalk plus base (TF0) which was buried in the membrane of the proton translocating vesicles . 4 . Vesicles capable of energy transformation were formed when TF1 came in contact with the surface of liposomes containing TF0 . On addition of phospholipids, the helix content of TF0 increased 3-fold . The role of F0 in forming channels for protons is discussed . 5 . The amino acid compositions of TF0, TF1, and TF0-F1 were compared . TF0 was not hydrophobic, despite its interaction with phospholipids . The phospholipid composition and other properties of the proton translocating vesicles were examined . Vesicles reconstituted from a mixture of phosphatidylethanolamine, phosphatidylgly-cerol, and cardiolipin in the same ratio as in the membranes had the highest activity. J Biochem (Tokyo), 1976 Jul, 80(1), 33 - 7 Enzymatic synthesis of oligonucleotides of defined sequence . The "single addition" of 2(3)-O-dihydrocinnamoyl-nucleoside 5'-diphosphate to a primer oligonucleotide catalyzed by a thermophilic polynucleotide phosphorylase; Kikuchi Y et al.; Several oligonucleotides of defined sequence were synthesized using 2'(3')-O-dihydrocinnamoyl-nucleoside 5'-diphosphates (DHC-NDP) as substrates for polynucleotide phosphorylase {EC 2.7.7.8} from Thermus thermophilus . The enzyme catalyzed the transfer of one nucleotidyl residue from each of the 2'(3')-O-dihydrocinnamoyl esters of CDP, UDP, and GDP to the 3'-terminus of the primer triadenosine diphosphate, (Ap)2A . The products were shown to be (Ap)3C, (Ap)3U, and (Ap)3G by enzymatic analysis. J Biochem (Tokyo), 1976 Jun, 79(6), 1331 - 44 The effect of temperature on ribose-5-phosphate isomerase from a mesophile, Thiobacillus thioparus, and a thermophile, Bacillus caldolyticus; Middaugh CR et al.; The enzyme ribose-5-phosphate isomerase {EC 5.3.1.6} was partially purified from a mesophilic organism, Thiobacillus thioparus, and from an extreme thermophile, Bacillus caldolyticus . The stability and kinetics of the two enzymes were compared with regard to temperature in the presence of a series of neutral salts and alcohols . The thermal stability of both enzymes was altered such that the salts (NH4)2SO4, NaCl, KCl, and LiCl increased stability, while LiBr, CaCl2, methanol, ethanol, and 1-propanol decreased stability . Ethylene glycol had little effect on the mesophilic enzyme, but increased the stability of the thermophilic protein . The kinetics of both enzymes were also affected by the salts and alcohols, and Arrhenius plots of two kinetic parameters, Km and Vmax, displayed discontinuities, or sharp changes in slope, at characteristic temperatures, TD . Neutral salts and alcohols altered the temperature of discontinuity in a sequence similar to that observed in studies of thermal stability . It is suggested that the slope change is due to temperature-dependent alterations in the enzymes at specific, but undefined, loci at the active site, although no evidence is offered for the absence of a larger conformation change in the entire enzyme. J Biochem (Tokyo), 1976 Jun, 79(6), 1245 - 52 Protein synthesis in a cell-free system from an extreme thermophile . Effects of preincubation in the cold on polyuridylic acid-dependent polyphenylalanine synthesis at high temperature; Ohno-Iwashita Y et al.; 1 . It was found that preincubation of the reaction mixture in the cold enhanced polyuridylic acid-directed polyphenylalanine synthesis by a cell-free extract of Thermus thermophilus HB8 at high temperature . 2 . The effect of preincubation was most marked at 10-25 degrees in the presence of 20 mM Mg2+ . Preincubation at 65 degrees failed to stimulate the incorporation . 3 . The presence of phenylalanyl-tRNA, polyuridylic acid, and ribosomes was essential for preincubation in the cold to be effective . 4 . A ternary complex of amino acyl-tRNA, polyuridylic acid, and a ribosome formed at low temperature was isolated by CPG-10 column chromatography; the isolated complex initiated polyphenylalanine synthesis effectively at high temperature . 5 . The amount of the ternary complex formed depends on the preincubation time and the concentration of Mg2+ . Since the amount of the complex correlated positively to the rate of polyphenylalanine synthesis at high temperature, the effectiveness of preincubation in the cold is presumably due to the formation of the ternary complex of phenylalanyl-tRNA, polyuridylic acid, and a ribosome. J S Afr Vet Assoc, 1976 Jun, 57(2), 101 - 4 {UHT--behandeling van melk (author's transl)}; Lombard SH; Pasteurisation of milk provides protection for the consumer against pathogens which may be present in the raw milk, and improves its keeping quality . Sterilisation provides indefinite keeping quality but has an undesirable effect on the flavour and nutritive value of the milk . Ultra high temperature (UHT) treatment produces a milk with prolonged shelf life at ambient temperatures yet has practically the same effect on colour, falvour and nutritive value as pasteurisation . UHT treatment of milk involves preheating to 80 degrees C and then quickly raising the temperature, either by indirect heating in a tubular heater or by direct steam injection, to 130 degrees-- to 150 degrees C at which temperature it is kept for 3 to 5 sec . Cooling follows immediately . Systems in operation in South Africa use steam injection for heating to sterilising temperatures . Evaporation cooling is obtained by subjecting milk to a partial vacuum . This removes any water added during heating by condensing steam and also removes steam-volatile off-flavours . UHT-treated milk is packed aseptically, usually into heat-sealed paperboard laminated cartons . Intact packages can be kept for up to three months . Absolute sterility cannot be obtained by UHT processing . The term "sterilising effect", introduced by Galesloot, means the log10 of the ratio of initial spore count to surviving spore count . A spoilage rate of not more than one litre package per 1000 is considered satisfactory . For laboratory control samples of the sterilized milk are incubated at temperatures favourable to germination of mesophilic and thermophilic spores respectively . After incubation the milk is examined for flavour and physical appearance, subjected to the standard plate count and tested for increase in acidity and decrease in stability towards the alcohol test . Milk for UHT treatment must possess protein stability in the alcohol test, be of good bacteriological quality, and a low spore count in particular. J Allergy Clin Immunol, 1976 Jun, 57(6), 518 - 24 Precipitating antibodies in a midwest dairy farming population toward the antigens associated with farmer's lung disease; Roberts RC et al.; A survey of the frequency of precipitins to the antigens of the thermophilic actinomycetes and Aspergillus species was conducted on serum samples from 1,045 farmers obtained at a 3-day exposition on modern farm equipment and farming practices in central Wisconsin . Each farmer filled out a questionnaire including socioeconomic information, lung disease history, exposure history, and smoking history . Precipitins were detected by the double-diffusion method . The antigen panel included eight thermophilic actinomycetes species and a mixture of Aspergillus species . Precipitins were found in the sera of 93 famers (8.9%) . The distribution of positive precipitins was: Micropolyspora faeni 63 (67.7%), Thermoactinomyces vulgaris 7 (7.5%), Thermomonospora viridis 2 (2.2%), M . faeni + T . viridis 16 (17.2%), M . faeni + T . vulgaris 1 (1.1%), Aspergillus species 4 (4.3%) . Of all the parameters tested for in the questionnaire, those with positive serology differed significantly from the whole population only in that a higher proportion of the positives reported exposure to silo gas and illness after uncapping silos . Comparison of the size of the farm and the number of dairy cows in the state of Wisconsin with the population samples indicated that the sample population was skewed toward those with larger farms and larger dairy herds . This study confirms that a significant proportion of the farm population in Wisconsin does have precipitins to the microorganisms associated with farmer's lung disease . Follow-up studies to establish the relationship between the positive precipitin reactions to the presence of clinical disease are now under way. J Biochem (Tokyo), 1976 Jun, 79(6), 1157 - 66 Active transport of alanine by thermostable membrane vesicles isolated from a thermophilic bacterium; Hirata H et al.; 1 . Thermostable membrane vesicles which were capable of active transport of alanine dependent on either respiration or an artificial membrane potential were isolated from the thermophilic aerobic bacterium PS3 . 2 . Uptake of alanine was dependent on the oxidation of ascorbate-phenazine methosulfate or on generated or exogenous NADH, but succinate and malate failed to drive the uptake . The optimum temperature for respiration-driven uptake of alanine was 45 to 60 degrees . 3 . Potassium ion-loaded vesicles were prepared by incubating vesicles at 55 degrees in 0.5 M potassium phosphate . The addition of valinomycin elicited rapid and transient uptake of alanine under the test conditions . Uptake of alanine in response to valinomycin was progressively enhanced by the addition of dicylohexylcarbodiimide, but was completely abolished in the presence of a proton conductor or synthetic permeable cation . The effect of dicyclohexylcarbodiimide was dependent on its concentration and was maximal at a concentration of 0.4 mM . 4 . The proton permeability of membrane vesicles was reduced by the addition of dicyclohexylcarbodiimide . A small but significant difference was found in the initial rates of proton uptake in the presence of dicyclohexylcarbodiimide with and without alanine . The results suggest that protons alanine are transported simultaneously in a stoichiometric ratio of 1 : 1 . 5 . The uptake of alanine was also driven by a pH gradient induced by an instantaneous pH drop in a suspension of alkali-loaded vesicles . Thus, alanine accumulation was driven not only by an electrical potential but also by a pH gradient . 6 . Addition of ATP resulted in the inhibition of alanine uptake dependent on artificial membrane potential . ATP hydrolysis by membrane ATPase created a membrane potential which was inside-positive, and this might decrease the effective membrane potential (generated by K+ efflux mediated by valinomycin) available to drive alanine uptake. Appl Environ Microbiol, 1976 Jun, 31(6), 819 - 25 Factors influencing the production of cellulases by Sporotrichum thermophile; Coutts AD et al.; Cellulase production and growth of a strain of Sporotrichum thermophile were studied by using a mineral salts medium supplemented with yeast extract and insoluble cellulose . The effects of cultural conditions, such as pH, nitrogen source, substrate concentration, and temperature, were examined . Maximum production of C1 and CX cellulases occurred at 45 C in 2 to 4 days, in the presence of 1% Solka/Floc as substrate, when NaNO3 or urea used as sources of nitrogen . Under these conditions, cellulolytic activity of culture filtrates appeared to be similar to that reported for Trichoderma viride grown in a favorable environment . However, comparable yields of cellulase were produced by S . thermophile in less than one-quarter the time required by mesophilic fungi. Appl Environ Microbiol, 1976 Jun, 31(6), 807 - 12 Production of extracellular alpha-glucosidase by a thermophilic Bacillus species; Suzuki Y et al.; Production of extracellular alpha-glucosidase was studied with strain KP 1006 of a new species of thermophilic Bacillus, which was isolated from soil samples by enrichment at 65 C . alpha-Glucosidase production was maximum at 60 C and at an initial pH of 6.5 . The final enzyme yield was increased by starch, maltose, glycerol, peptone, and yeast extract but reduced by acetate and gluconate, alpha-Glucosidase was formed in the cytoplasm and accumulated as a large pool during the logarithmic growth phase . At a midpoint of this period, the enzyme appeared in the culture broth, and its level increased until the end of the stationary phase. Can J Microbiol, 1976 Jun, 22(6), 817 - 25 beta-Galactosidase from Bacillus stearothermophilus; Goodman RE et al.; Several strains of thermophilic aerobic spore-forming bacilli synthesize beta-galactosidase (EC 3.2.1.23) constitutively . The constitutivity is apparently not the result of a temperature-sensitive repressor . The beta-galactosidase from one strain, investigated in cell-free extracts, has a pH optimum between 6.0 and 6.4 and a very sharp pH dependence on the acid side of its optimum . The optimum temperature for this enzyme is 65 degrees C and the Arrhenius activation energy is about 24 kcal/mol below 47 degrees C and 16 kcal/mol above that temperature . At 55 degrees C the Km is 0.11 M for lactose and 9.8 X 10(-3) M for 9-nitrophenyl-beta-D-galactopyranoside . The enzyme is strongly product-inhibited by galactose (Ki equals 2.5 X 10(-3) M) . It is relatively stable at 50 degrees C, losing only half of its activity after 20 days at this temperature . At 60 degrees C more than 60% of the activity is lost in 10 min . However, the enzyme is protected somewhat against thermal inactivation by protein, and in the presence of 4 mg/ml of bovine serum albumin the enzyme is only 18% inactivated in 10 min at 60 degrees C . Its molecular weight, estimated by disc gel electrophoresis, is 215 000. J Biol Chem, 1976 May 25, 251(10), 3122 - 7 The primary structure of Bacillus subtilis and Bacillus stearothermophilus 5 S ribonucleic acids; Marotta CA et al.; The nucleotide of Bacillus stearothermophilus 5 S RNA is pC-C-U-A-G-U-G-A-C-A-A-U-A-G-C-G-(G-A-G-A-G-G-)-A-A-A-C-A-C-C-C-G-U-U-C-C-C-A-U-C-C-C-G-A-A-C-A-C-G-G-A-A-G-U-U-A-A-G-C-U-C-U-C-C-A-G-C-G-C-C-G-A-U-G-G-U-A-G-U-U-G-G-G-G-C-C-A-G-C-G-C-C-C-C-U-G-C-A-A-G-A-G-U-A-G-G-U-C-G-U-U-G-C-U-A-G-G-COH; the nucelotide sequence of Bacillus subtilis 5 S RNA is pU-U-U-G-G-U-G-G-C-G-A-U-A-G-C-G-A-A-G-A-G-G-U-C-A-C-A-C-C-C-G-U-U-C-C-C-A-U-A-C-C-G-A-A-C-A-C-G-G-A-A-G-U-U-A-A-G-C-U-C-U-U-C-A-G-C-G-C-C-G-A-U-G-G-U-A-G-U-C-G-G-G-G-G-U-U-U-C-C-C-C-C-U-G-U-G-A-G-A-G-U-A-G-G-A-C-G-C-C-G-C-C-A-A-G-COH . Comparison of the sequence of B . stearothermophilus 5 S RNA to the sequence of B . subtilis 5 S RNA and to that of Bacillus megaterium 5 S RNA (Pribula, C . D., Fox, G . E., Woese, C . R., Sogin, M., and Pace, N . (1974) FEBS Lett . 44, 322-323) indicates that the 5 S RNA isolated from the thermophile contains unique nucleotide sequences not found in 5 S RNAs isolated from the two mesophilic species of genus Bacillus. J Biol Chem, 1976 May 25, 251(10), 3134 - 9 Purification and characterization of a repressible alkaline phosphatase from Thermus aquaticus; Yeh MF et al.; A repressible alkaline phosphatase has been isolated from the extreme bacterial thermophile, Thermus aquaticus . The enzyme can be derepressed more than 1,000-fold by starving the cells for phosphate . In derepressed cells, nearly 6% of the total protein in a cell-free enzyme preparation is alkaline phosphatase . The enzyme was purified to homogeneity as judged by disc acrylamide electrophoresis and sodium dodecyl sulfate electrophoresis . By sucrose gradient centrifugation it was established that the enzyme has an approximate molecular weight of 143,000 and consists of three subunits, each with a molecular weight of 51,000 . Tris buffer stimulates the activity of the enzyme, which has a pH optimum of 9.2 . The enzyme has a broad temperature range with an optimum of 75-80 degrees . The enzyme catalyzes the hydrolysis of a wide variety of phosphorylated compounds as do many of the mesophilic alkaline phosphatases . The Michaelis constant(Km) for the enzyme is 8.0 X 10(-4) M . Amino acid analysis of the protein revealed little in the amino acid composition to separate it from other mesophilic enzymes which have been previously studied. Mikrobiologiia, 1976 May-Jun, 45, 460 - 5 {Production of protease during continuous cultivation of the thermophilic bacterium Bacillus brevis}; Loginova LG et al.; Conditions for continuous cultivation of the thermophilic strain of Bacillus brevis prodlcing protease are substantiated . The culture was cultivated in order to obtain proteolytic enzymes taking into consideration data concerning kinetics of the bacterial growth and enzyme synthesis, as well as results of studying accumulation of the pool of mRNA specific of protease synthesis, The rates of dilution and flow (approximately 0.4 hr-1) have been found which provide a high specific productivity of the culture. J Gen Microbiol, 1976 May, 94(1), 131 - 41 Regulatory properties of an inducible aliphatic amidase in a thermophilic bacillus; Thalenfeld B et al.; A thermophilic bacillus growing on acetamide as both carbon and nitrogen sources produces an inducible amidase . This amidase hydrolysed the following amides in decreasing order or activity, in comparison with acetamide (1.00): propionamide (0.97), fluoroacetamide (0.84), formamide (0.35) and glycinamide (0.12) . Cyanoacetamide, dimethylacetamide, dimethylformamide and urea also induced the synthesis of the amidase, but were not substrates of the enzyme . Studies with protoplasts suggest that the amidase is located in the cytoplasm . Glucose strongly inhibited amidase synthesis; and limiting nitrogen did not release this inhibition . Urea strongly inhibited amidase activity in a competitive manner; but the inhibition caused by iodoacetamide and cyanoacetamide was non-competitive . Both thioacetamide and thiourea were effective inhibitors of enzyme induction . Bacteria grown on a succinate-minimal medium exhibited a lag in amidase synthesis, which could be eliminated by decreasing the concentration of succinate . Acetate- or pyruvate-grown cultures behaved similarly, while those grown on alanine or glutamate exhibited no lag in enzyme induction . In the mutant strain E21, repression of amidase synthesis by glucose was much less evident and no lag for induction was apparent with any of the other carbon sources mentioned. Rev Asoc Argent Microbiol, 1976 May-Aug, 8(2), 74 - 81 {Regulation of citrate synthese in bacteria: Comparison of the action of various effectors on the enzymes of Rhodospirillum rurbum and Bacillus stearothermophilus}; Higa AI et al.; A comparative study of the citrate synthases purified from the facultatively photosynthetic bacterium Rhodospirillum rubrum (Gram negative) and the thermophile Bacillus stearothermophilus (Gram positive) was made . The citrate synthase from R . rubrum was activated by KCl (6-fold at 0.1 M KCl) and, less effectively, by NaCl and NH4Cl . Its molecular weight was about 300,000 . The enzyme was strongly inhibited by NADH, and this inhibition was counteracted by AMP . The citrate synthase from B . stearothermophilus was little affected by KCl, NaCl and NH4Cl, all of which activated by about 25% at 0.1 M . Its molecular weight was ca 100,000 . The enzyme was not affected by NADH or AMP . Both citrate synthases were insensitive to alpah-oxoglutarate concentrations up to 5 mM, and were inhibited by ATP; the B . stearothermophilus enzyme was more strongly inhibited than the R . rubrum enzyme . In both cases the ATP inhibition was strictly competitive towards acetyl-CoA and non-competitive towards oxaloacetate . Both enzymes, in spite of the peculiar physiological properties of their bacterial sources, followed the close correlation between the properties of the citrate synthase and the taxonomical position of the microorganism, proposed by Weitzman and his co-workers. J Allergy Clin Immunol, 1976 May, 57(5), 417 - 21 Immunologic cross-reactions among thermophilic actinomycetes associated with hypersensitivity pneumonitis; Kurup VP et al.; Antibodies produced in rabbits against Micropolyspora faeni, Thermoactinomyces vulgaris, T . sacchari, Thermoactinomyces candidus, and Saccharomonospora viridis were tested against antigens derived from many strains of thermophilic actinomycetes for precipitating antibodies by immunodiffusion test . It was found that immune sera reacted strongly against antigens from strains belonging to the same species and weakly against antigens from different species of thermophilic actinomycetes . However, sera from farmer's lung patients showed cross-reactivity against antigens from different species . This may be because the patient is sensitized to multiple species of thermophilic actinomycetes present in the environment and developed antibodies against most of them. Mikrobiologiia, 1976 May-Jun, 45, 417 - 9 {Alternativity of methane assimilation pathways in obligate methylotrophs}; Shishkina VN et al.; The activity of key enzymes involved in the primary pathways of methane assimilation and enzymes of the citrate cycle was determined in various obligate methylotrophs: mesophilic, thermotolerant, and thermophilic . The bacteria are characterized by the membrane ultrastructure of the I type, high activity of hexosephosphate synthase, NAD- and NADP-specific isocitrate dehydrogenase, and the absence of alpha-ketoglutarate dehydrogenase . The bacteria also displayed the activity of key enzymes of the serine cycle, hydroxypyruvate reductase, and serineglyoxylate aminotransferase . Therefore, both the ribulose monophosphate and serine pathways are involved in methane assimilation, and the division of methanotrophs into two groups, according to their metabolism, is tentative. Eur J Biochem, 1976 May 1, 64(2), 341 - 50 Alcohol oxidases of Kloeckera sp . and Hansenula polymorpha . Catalytic properties and subunit structures; Kato N et al.; 1 . Alcohol oxidase (alcohol: oxygen oxidoreductase) of a thermophilic methanol-utilizing yeast, Hansenula polymorpha DL-1, was isolated in crystalline form . 2 . This alcohol oxidase of H . polymorpha was more stable to heat than was the enzyme of Kloeckera sp . This difference in heat stability is compatible with the difference in growth temperatures for both yeasts . 3 . The crystalline alcohol oxidases of both yeast oxidized the lower primary alcohols (C-2 to C-4) as well as methanol . The apparent Km values for the methanol of Kloeckera and H . polymorpha enzymes were 0.44 and 0.23 mM, respectively . The enzymes could also oxidize formaldehyde to formate, and were inactivated by relatively low concentrations of hydrogen peroxide . 4 . The molecular weight for both enzymes was calculated to be about 670000 . Each enzyme is composed of eight identical subunits (molecular weight 83000) and contains eight moles of FAD as the prosthetic group . The NH2-terminal and COOH-terminal amino acids of H . polymorpha enzyme were identified as alanine and phenylalanine, respectively . The octameric subunits model of each enzyme was confirmed by electron micrographs, which showed an octad aggregate, composed of two tetragons face to face. Biochemistry, 1976 Apr 20, 15(8), 1749 - 55 Glutamine synthetase of Bacillus stearothermophilus . Regulation, site interactions, and functional information; Wedler FC et al.; The action of various feedback modifiers on Bacillus stearothermophilus glutamine synthetase has been investigated by initial velocity kinetics, using the Mn2+-stimulated biosynthetic assay at 55 degrees C . The most potent inhibitors, used singly, are AMP, L-glutamine, and L-alanine . Other modifiers of significance include glycine, CTP, L-histidine, glucosamine 6-phosphate, and GDP . Marked synergism of action is observed for AMP in the presence of L-glutamine, L-histidine, ADP, or glucosamine 6-phosphate (glucosamine-6-P), and for CTP with ADP or GDP . Inhibition by saturating levels of many modifiers is either less than 100%, or is not overcome by elevated substrate levels, or both . This argues for modifier binding sites separate from substrate sites, notably in the cases of AMP, L-glutamine, glycine, L-alanine, glucosamine-6-P, and CTP . Glycine and L-alanine are Vmax inhibitors, whereas L-glutamine, glucosamine-6-P, GDP, and CTP alter the binding of L-glutamate . ADP and L-histidine apparently can compete directly with MnATP, but AMP alters Mn-ATP binding from a separate site . The action of several modifiers requires or is enhanced by bound substrates . Considerable antagonistic interaction is observed in experiments with modifier pairs, but the most potent inhibitors show synergistic or cumulative (independent) interactions . One may interpret antagonistic effects as due to (a) overlapping modifier domains, or (b) separate but antagonistically interacting sites . Either interpretation leads to a scheme for modifier-substrate and modifier-modifier site interactions in which the thermophilic enzyme must maintain and stabilize a great deal of complex functional information under extreme environmental conditions. Biochemistry, 1976 Apr 20, 15(8), 1692 - 6 DNA-dependent RNA polymerase from the thermophilic bacterium Caldariella acidophila . Purification and basic properties of the enzyme; Cacace MG et al.; A DNA-dependent RNA polymerase has been isolated from Caldariella acidophila, a thermophilic bacterium living in acidic hot springs at temperatures ranging from 63 to 89 degrees C . The enzyme was purified 180-fold and is composed of five different subunits having the following molecular weights: a = 127000, b = 120000, c = 72000, d = 65000, and e = 38000 . The enzyme is activated by Mn2+ and Mg2+ and exhibits optimal activity in the presence of 0.5 mM Mn2+ . The activity depends on ionic strength, with a maximum at 0.25 M KCl, and exhibits a pH optimum at 7.8 in the presence of Tris-HCl buffer . The enzyme shows a high degree of thermophilicity, its temperature optimum being 80 degrees C in the in vitro assay . The thermophilicity of C . acidophila RNA polymerase allows studies on enzyme-template interactions to be performed in a temperature range where many templates are close to their Tm. Eur J Biochem, 1976 Apr 15, 64(1), 57 - 68 Purification and properties of D-glyceraldehyde-3-phosphate dehydrogenase from an extreme thermophile, Thermus thermophilus strain HB 8; Fujita SC et al.; 1 . D-Glyceraldehyde-3-phosphate dehydrogenase from an extreme thermophile, T . thermophilus strain HB8, was purified and crystallized . 2 . The enzyme was found to possess remarkable heat stability, being slowly inactivated at 90 degrees C . 3 . Basic kinetic constants and pH profile are reported . The enzyme was activated 25-fold by 90 mM NH4Cl, and also by ethanol up to 5-fold at 30 degrees C . 4 . The enzyme was found to be far more resistant to urea or sodium dodecylsulfate than the rabbit enzyme . 5 . The enzyme was shown to be a tetramer of molecular weight 130000--135000 . Amino acid composition analysis revealed no unusual features . Circular dichroic spectra suggested that the contents of the ordered structure of the thermophile enzyme are similar to those of the rabbit enzyme . 6 . The other catalytic properties of the thermophile enzyme are discussed in comparison with those of the enzymes from other sources. J Mol Evol, 1976 Apr 9, 7(3), 197 - 213 A comparison of the 16S ribosomal RNAs from mesophilic and thermophilic bacilli: some modifications in the Sanger method for RNA sequencing; Woese C et al.; Two modifications in the Sanger two dimensional electrophoretic procedure for RNA analysis are reported . One increases resolution on the primary fingerprint to the point that digests of large RNAs, of the size 1500-3000 nucleotides yield well resolved fingerprint patterns . The other is a novel endonucleolytic procedure that proves useful in determining sequences of the large oligonucleotides produced by T1 ribonuclease . These modifications have been used in determining the catalogs of oligomers produced by T1 ribonuclease digestion of 16S rRNAs from three related organisms, Bacillus subtilis, B.pumilus and B.stearothermophilus . The possible effects of adaptation to a thermophilic niche on ribosomal RNA primary structure and the phylogenetic relatedness of the two mesophilic Bacilli are discussed. Ann Intern Med, 1976 Apr, 84(4), 406 - 13 Interstital lung disease due to contamination of forced air systems; Fink JN et al.; Eight patients had hypersensitivity pneumonitis due to contaminated home or office forced-air heating or air-conditioning systems . We studied their clinical and laboratory features, and the results indicated that this disease may occur as an acute or insidious form differing in type and intensity of respiratory and systemic symptoms . Thermophilic actinomycetes contaminatinf the forced air systems were identified as the sensitizing agents in most cases . Precipitating antibodies to the organisms could be shown in the serums of the patients and the antigen identified by immunofluorescent studies in the three lung biopsies examined by this method.Inhalation challenge studies with the cultured organism or other materials obtained from the forced air systems reproduced the clinical syndrome in the four patients tested . Avoidance of the contaminated system, and the use of corticosteroids in more severe cases,seems to be appropriate therapy for patients with this disease. Ann Med Interne (Paris), 1976 Apr, 127(4), 295 - 307 {Pulmonary reactions to non-infective antigenic contaminants}; Akoun G et al.; There are different categories of non-infectious antigenic contaminants capable of provoking pulmonary reactions: thermophilic actinomycetales, fungi, vegetable particles, protein antigens, arthropods, drugs; their identification is based on their isolation, culture, and the study of their biochemical composition . The pulmonary reactions depend on the size and number of antigenic particles inhaled and by the underlying background . The immunological mechanisms brought into play a complex Arthus's phenomenon: in addition to the IgGs, the IgEs and the different factors of complement intervene; the recent acquisitions of immunomorphology enable better comprehension of the histogenesis of the pulmonary granuloma . The diagnosis of extrinsic allergic alveolitis is founded on well defined criteria of clinical, radiological, respiratory functional, immunological and histophthological order . These conditions are met with in agricultural, industrial and urban environments or are related to iatrogenic diseases . Many problems still remain obscure with regards to the pathogenesis and integration within the framework of immunological pneumonia of various conditions, and especially certain sarcoidosis. Arch Microbiol, 1976 Mar 19, 107(2), 223 - 4 {Thielavia heterothallica spec . nov . (Abb.1) Status conidialis: Chrysosporium thermophilum (Apinis) von Klopotek}; von Klopotek A; Thielavia heterothallica spec . nov . is described as a heterothallic, thermophilic fungus with spherical, black, non-ostiolate cleistothecia; elliposidal evanescent asci which contain eight one-celled ellipsoidal ascospores, darkening to deep brown to black, with one germ pore . The conidial state is Chrysosporium thermophilum (Apinis) von Klopotek. J Bacteriol, 1976 Mar, 125(3), 1211 - 3 Occurrence of phosphenolpyruvate carboxylase in the extremely thermophilic bacterium Thermus aquaticus; Bridger GP et al.; In the extreme thermophile Thermus aquaticus, phosphoenolpyruvate carboxylase catalyzes carbon dioxide fixation on the C3 metabolite phosphoenolpyruvate, producing oxaloacetate . In a moderately thermophilic Bacillus species this function is fulfilled by pyruvate carboyxlase . Like several of its mesophilic counterparts, the Thermus enzyme exhibits a requirement for acetyl coenzyme A. Mikrobiologiia, 1976 Mar-Apr, 45(2), 284 - 90 {The physiology of thermophilic and mesophilic bacilli during development at optimal and submaximal temperatures}; Pozmogova IN et al.; Thermophilic bacilli, contrary to mesophilic, grow equally well under aerobic and anaerobic conditions . This may be due to the presence of more active anaerobic dehydrogenases in thermophilic organisms . The activity of glucose-6-phosphate dehydrogenase and alcohol dehydrogenase, and the content of ATP, increased in the cells of thermophilic Bac . stearothermophilus 159 and mesophilic Bac . megaterium M strains growing at temperatures close to maximal . The activity of glutamate dehydrogenase under these conditions was inhibited. J Bacteriol, 1976 Mar, 125(3), 850 - 4 Thermal denaturation of mesophilic and thermophilic 5S ribonucleic acids; Varricchio F et al.; The Tm of Bacillus stearothermophilus 5S ribonucleic acid (RNA) is 1.5 +/- 0.5 C higher than that of 5S RNAs from B . subtilis and Escherichia coli . Melting in 50% methanol and in formaldehyde indicate that both base stacking and helical regions are involved in the slightly increased thermal stability of B . stearothermophilus 5S RNA . It is probable that the 5S RNA makes only a minor contribution to the thermostability of B . stearothermophilus 50S ribosomal subunits. J Biochem (Tokyo), 1976 Mar, 79(3), 469 - 77 Studies on alpha-amylase from a thermophilic bacterium . II . Thermal stability of the thermophilic alpha-amylase; Hasegawa A et al.; The effect of pH, mental ions, and denaturing reagents on the thermal stability of thermophilic alpha-amylase {EC 3.2.1.1} were examined . The enzyme was most stable at around pH 9.2, which is coincident with the isoelectric point of the enzyme . The stability of the enzyme was increased by the addition of calcium, strontium, and sodium ions . The addition of calcium ions markedly stabilized the enzyme . The protective effects of calcium and sodium ions were additive . At room temperature, no detectable destruction of the helical structure of the enzyme was observed after incubation for 1 hr in the presence of 1% sodium dodecylsulfate, 8 M urea or 6 M guanidine-HC1 . The addition of 8 M urea or 6 M guanidine-HC1 lowered the thermal denaturation temperature of the enzyme . The enzyme contained one atom of tightly bound intrinsic calcium per molecule which could not be removed by electrodialysis unless the enzyme was denatured . The rate constants of inactivation and denaturation reactions in the absence and presence of calcium ions were measured and thermodynamic parameters were determined . The presence of calcium ions caused a remarkable decrease in the activation entropy. Biochemistry, 1976 Feb 24, 15(4), 842 - 8 Purification and properties of the thermostable acid protease of Penicillium duponti; Emi S et al.; An acid protease produced by the thermophilic fungus Penicillium duponti K 1014 has been purified by consecutive ion-exchange and gel permeation chromatography, and crystallized from aqueous acetone solution . The purified endopeptidase gave a symmetrical schlieren peak by sedimentation velocity, and was found to be homogeneous upon disc gel electrophoresis at pH 9.5 . The enzyme was most active at pH 2.5 against milk casein and showed high thermostability . An isoelectric point of 3.81 was found by isoelectric focusing . A minimum molecular weight of 41 590 was calculated from the amino acid composition, adopting an arginine content of one residue per mole of enzyme . This minimum molecular weight is in good agreement with the value of 41 000 previously found by gel permeation (Hashimoto, H., Iwaasa, T., and Yokotsuka, T . (1973), Appl . Microbiol . 25, 578) . Besides the thermostability, the purified P . duponti protease differs from other well-characterized acid proteases in that it contains carbohydrate, 4.33% expressed as glucose . The enzyme was not affected by p-bromophenacyl bromide, but was completely inactivated by alpha-diazo-p-bromoacetophenone, diazoacetyl-DL-norleucine methyl ester, and diazoacetylglycine ethyl ester, in the presence of Cu2+ . The complete inactivation of the protease by diazoacetyl-DL-norleucine methyl ester resulted in the specific incorporation of 1 mol of norleucine/mol of enzyme . On the basis of similar behavior of other acid proteases toward this inactivator, the results suggest the presence at the active site of an unusually reactive carboxyl group, involved in the catalytic function . The naturally occurring pepsin inhibitor of Streptomyces naniwaensis {Murao, S., and Satoi, S . (1970), Agric . Biol . Chem . 34, 1265} inhibited also the protease, at a threefold molar excess with respect to the enzyme. Arch Microbiol, 1976 Feb, 107(1), 49 - 55 Altered phospholipid metabolism in a temperature-sensitive mutant of a thermophilic bacillus; Kostiw LL et al.; The phospholipid metabolism of a temperature-sensitive mutant of a thermophilic bacillus was studied after the shift from a permissive (58 degrees C) to a restrictive (65 degrees C) growth temperature . During the short period of growth of the mutant at 65 degrees C, the proportions of cardiolipin and its 3-acyl derivative (lyso-cardiolipin) increased, and the proportions of phosphatidylglycerol and phosphatidylethanolamine decreased on cell dry weight basis . In 32P incorporation and turnover experiments, phosphatidylglycerol showed the most rapid uptake and loss of the label . Turnover of cardiolipin, limited to a short period, ceased 18 min after the shift, as did the turnover of phosphatidylethanolamine . In the absence of net phospholipid synthesis, there was a quantitative conversion of phosphatidylglycerol to cardiolipin and an increase in the proportion of lyso-cardiolipin . Chloramphenicol, added to the medium at the time of the shift, reduced the rate of phospholipid synthesis, prevented the increase in the proportions of cardiolipin and lyso-cardiolipin, and slowed the decrease in the proportions of the other two phospholipids . The results indicated a defect in the regulatory mechanism(s) of phospholipid metabolism in the mutant at the restrictive temperature. J Allergy Clin Immunol, 1976 Feb, 57(2), 174 - 6 Home humidifier thermophilic actinomycete isolates; Seabury J et al.; Twenty-seven thermophilic actinomycete isolates obtained from humidification systems or living-bedroom areas of individuals with suspected but unproved home environment-related respiratory disease were characterized morphologically and biochemically . All isolates were demonstrated to be members of the Thermoactinomyces genus . Two previously misidentified isolates were reclassified as a Thermoacetinomyces sp . Thus, all of our thermophilic actinomycete humidifier isolates studied to date have been identified as either Thermoactinomyces vulgaris or Thermoactinomyces sp . Large numbers of unidentified thermotolerant bacteria have also been isolated from virtually all residual humidifier water samples and their possible role in production of "humidifier-associated" respiratory disease is unknown. Arch Microbiol, 1976 Feb, 107(1), 75 - 80 Effect of temperature on the viability of Bacillus stearothermophilus; Hashizume S et al.; One of the obligate thermophilic bacteria, Bacillus stearothermophilus, was unable to grow at temperatures below 35 degrees C . About 80% of the population in the bacterial culture died at the temperatures, and the same extent of loss in either of the activities of oxygen consumption or synthesis of protein or nucleic acid of the organisms was observed . With the progress of death of the organisms, reduced nicotinamide-adenine dinucleotide came to be oxidized by the organisms, enzymes such as fructose-1,6-diphosphate aldolase, when the organisms were washed with phosphate buffer, were leaked out of the organisms, and an increasing amount of ribonucleoprotein was released into the culture medium . The change of the membrane state was then suggested to be one of the possible causes for the death of the organisms at the temperatures. Can J Microbiol, 1976 Feb, 22(2), 165 - 76 Production, purification and characterization of thermomycolase, the extracellular serine protease of the thermophilic fungus Malbranchea pulchella var . sulfurea; Ong PS et al.; The thermophilic fungus Malbranchea pulchella produces a single extracellular, alkaline, serine protease when grown at 45 degrees C, on 2% casein as sole carbon source . The growth-associated production of protease in submerged cultures was inhibited by addition of glucose, amino acids, or yeast extract . A simple four-step purification which yields homogeneous protease in 78% yield is described . The protease has an isoelectric point of 6.0, a pH optimum of 8.5, and is completely inhibited by serine protease inhibitors . A specificity study with small synthetic ester substrates indicated that the protease preferentially hydrolyzed bonds situated on the carboxyl side of aromatic or apolar amino acid residues which are not beta-branched, positively charged or of the D configuration . Peptidase substrates and others such as N-acetyl-L-tyrosine-ethyl ester were not hydrolyzed . The protease was stable over a broad range of pH (6.5-9.5 at 30 degrees C, 20 h), and was particularly thermostable (t1/2 = 110 min at 73 degrees C, pH 7.4) in the presence of Ca2+ (10 mM) . Macromolecules and Ca2+ also provide protection against the significant autolysis which occurs at pure protease concentrations greater than 0.01 mg/mo, as well as against surface denaturation which is enhanced by the presence of a silicone antifoam agent . Hence the stability of protease in submerged cultures is rationalized. J Dairy Sci, 1976 Feb, 59(2), 200 - 2 Microorganisms and characteristics of laban; Baroudi AA et al.; Laban had a titratable acidity of about 1.0%, a pH of 4.25, an ethanol content of 1.25%, and contained 4.2 mug acetaldehyde and 34 mug acetoin/ml . There was no diacetyl . Five microorganisms, classified as Streptococcus thermophilus, Lactobacillus acidophilus, Leuconostoc lactis, Kluyveromyces fragilis, and Saccharomyces cerevisiae, were responsible for the fermentation . Streptococcus thermophilus and L . acidophilus were responsible for acid production with S . thermophilus producing acid more rapidly . Most of the acetaldehyde was produced by K . fragilis, little ethanol was found in absence of S . cerevisiae, and the acetoin was producted by S . thermophilus. Clin Allergy, 1976 Jan, 6(1), 19 - 25 A quantitative study on the activation of the alternative pathway of complement by mouldy hay dust and thermophilic actinomycetes; Edwards JH; Materials associated with the induction of farmer's lung were incubated with fresh normal human serum in the presence of magnesium ethylene glycol tetra-acetic acid (MgEGTA) or ethylene diamine tetra-acetic acid (EDTA) and results compared with material known to activate the alternative pathway of complement--zymosan . Results show that Micropolyspora faeni organisms are as active as zymosan in reducing complement (C) levels in the presence of MgEGTA, with a 50% reduction in CH50 at approximately 140 mug/ml . Thermoactinomyces vulgaris organisms produced a 50% CH50 reduction at approximately 1-25 mg/ml and two samples of respirable mouldy hay dust (MHD) at approximately 5-6 mg/ml whereas extracts of M . faeni and T . vulgaris reduced the CH50 titre by 17% and 39% respectively at 16 mg/ml in the presence of MgEGTA . Organisms and extracts did not reduce the CH50 titre in the presence of EDTA even at the maximum concentration quoted by more than 3%, thus it is considered that alternative pathway activation was responsible for C utilization in the presence of MgEGTA . Respirable MHD used less than 4% available C at 4 mg/ml in the presence of EDTA but at 8 mg/ml dust 11% and 28% available CH50 were used compared with 79% and 81% respectively in the presence of MgEGTA . The elution of immunoglobulin binding material from MHD may be responsible for apparent CH50 consumption in the presence of EDTA. Folia Parasitol (Praha), 1976, 23(4), 327 - 42 Influence of microclimate on the life cycle of the common tick Ixodes ricinus (L.) in thermophilic oak forest; Daniel M et al.; Under conditions of the South Moravian thermophilic oak forest (Valtice near Breclav), the life cycle of the common tick Ixodes ricinus was studied by continuous recording of main elements of microclimate (temperature and humidity) in three different biotopes: forest, margin of the forest and meadow . Simultaneously conditions and the process of tick hibernation were studied in four soil layers (surface, depths of 10, 20 and 30 cm) . Observations made in the winter and vegetation periods were assessed by mathematical-statistical tests . Results obtained in the forest biotope and at its margin are presented in this paper; results from the meadow biotope will be published separately. J Biochem (Tokyo), 1976 Jan, 79(1), 35 - 42 Studies on an alpha-amylase from a thermophilic bacterium . I . Purification and characterization; Hasegawa A et al.; An amylase-producing thermophilic bacterium was isolated from a Japanese hot spring . The thermophilic cells were gram-negative, nonsporulating, nonpigmented rods and were motile with flagella . Alpha-Amylase {EC 3.2.1.1) purified from the thermophile was studied . The enzyme was more heat-stable than the corresponding enzymes from mesophilic sources, and 50% loss of activity was observed after incubation for 1 hr at 90 degrees . The thermophilic enzyme resembled the corresponding mesophilic enzymes in many respects, such as kinetic parameters, physicochemical properties, and amino acid composition . The molecular weight of the enzyme was 50,000 and the enzyme contained 2 moles of half-cystine residues per mole of protein, but there were no disulfide crosslinkages . The alpha-helix content of the enzyme was estimated to be about 20% from CD spectra, a value similar to those of other alpha-amylases . The isoelectric point of the enzyme was at pH 9.2. Experientia Suppl, 1976, 26, 375 - 89 Thermophilic and mesophilic enzymes from B . caldotenax and B . stearothermophilus: properties, relationships and formation; Frank G et al.; 1) The adaptive system of thermophilic bacteria, as demonstrated with B . caldotenax, seems to be suitable to produce thermophilic and mesophilic enzymes for comparative studies . 2) If it may be assumed that the extensive homologies in the N-terminal sequences of the LDHs also extend over the entire polypeptide chain, comparison of these sequences together with investigation on the 3-dimensional structure offer the possibility of elucidating those structural details which may be responsible for thermostability and the other thermophilic properties . However, the difficulty still remains that the latter may be obscured by differences not related to thermostability etc . Neverthless it may be hoped that comparison of the full sequences of not only the LDHs but also of a sufficient number of other enzymes of the same system will yield such details . 3) A further interesting goal with respect to the mechanism of enzyme adaptation would be reached if the differences in amino acid sequence of thermophilic and mesophilic LDH enzymes would throw light on the type of the amino acids always being exchanged . Here from the very hypothetical point of view the question arises as to whether the bacterial cell during the metabolic adaptation process or even by mutation/selection is able to modify just those few amino acid residues thermodynamically important for thermostability . Alternatively: does there exist a "rule" by which certain amino acid residues are invariably exchanged on a change for thermophilic to mesophilic enzymes? 4) Problems not mentioned here arise with B . stearothermophilus, which can be adapted poorly via the spores or on intermediate temperatures . Of great importance, but also a special problem in these studies on thermophilic and mesophilic enzymes produced by the same bacterium are a) the characterization of the thermophilic (70 degrees or 55 degrees) and mesophilic (37 degrees) bacterial variants (in respect to type), b) the control of homogeneity of the bacterial culture (contamination, mixed population), c) proof of the genetic identity of the 70 degrees- (55 degrees-) and 37 degrees -variant of B . caldotenax and B . stearothermophilus, which differ greatly in their phenotypes, for example in their metabolism, cell- or colony merphology . The taxonomical-biochemical identity or also the identity of morphology of the sporangia, since this should be an expression of the temperature dependent phenotype, cannot be used unconditionally as criteria of identity . Criteria such as the presence of identical enzymes in both variants or the identity of the genome (use of genetic markers, anlaysis of the DNA) are more reliable . Experiments with both variants of B . caldotenax demonstrated an identically high content of cytosine plus guanine in their DNA: 62.2% in the thermophilic DNA and 66.8% in the mesophilic DNA . In the thermophilic B . stearothermophilus the C+G content of the DNA was 56.5% and in the mesophilic variant 57.1%... Experientia Suppl, 1976, 26, 249 - 62 Chemical, physical and enzymatic comparisons of formyltetrahydrofolate synthetases from thermo- and mesophilic Clostridia; O'brien WE et al.; Formyltetrahydrofolate synthetase from the mesophiles Clostridium cylindrosporum, C . acidiurici, and C . formicoaceticum and the thermophile C . thermoaceticum have been purified to homogeneity, the first two by Rabinowitz and Himes and their coworkers and the latter two by ourselves . The physical properties of these proteins are very similar . All four enzymes are tetrameric and all are activated by NH4+ or K+, and the mechanism of this activation always involves a decrease in Km for formate . The enzyme from C . formicoaceticum is more thermostable and has a higher temperature optimum than the C . cylindrosporum or C . acidiurici enzymes and the C . thermoaceticum enzyme is the most thermostable . Comparisons of the amino acid compositions indicate possible correlations between thermostability and an increased content of isoleucine, arginine and tryptophan and a decreased content of phenylalanine and aspartic acid . Both of the less thermostable enzymes dissociative above 37 degrees if NH4+ or K+ are removed, but neither of the more stable enzymes do . In 25% D2O, the C . formicoaceticum enzyme is destabilized at elevated temperatures, relative to the C . thermoaceticum enzyme . It is possible that stronger hydrophobic interactions between subunits are responsible for increased thermostability in these enzymes. Experientia Suppl, 1976, 26, 207 - 22 Purification and catalytic properties of "thermostable" fumarase from Bacillus stearothermophilus NU-10 and Thermus X-1; Cook WR et al.; Fumarase (L-malate hydro-lysase E.C.4.2.1.2) was purified from the thermophilic bacteria Bacillus stearothermophilus NU-10 (optimum growth temperature 62-63 degrees C) and Thermus X-1 (optimum growth temperature 70 degrees C) . The furmarase from Thermus X-1 is slightly more thermostable and has an "optimum" catalytic reaction temperature of 83 degrees C as compared to 81 degrees C for the B . stearothermophilus enzyme . Increased thermostability of these fumarases permitted an examination of the properties of the enzyme catalyzed reaction at temperatures higher than had previously been possible with the furmarases from mesophilic bacteria or higher plant and animal sources . Beyong the observed thermostability of the thermophilic fumarases, the catalytic properties of thermophilic fumarases were very similar to those observed with bacterial fumarase or the well characterized pig heart fumarase (effect of temperature on substrate affinities, pH optimum, substrate inhibition by fumarate, and Haldane relationship) . These similarities suggest that thermophilic enzymes may be useful in the general study of enzyme reaction mechanisms. Ontogenez, 1976, 7(1), 93 - 5 {Heat resistance of aldolase of the hybrid embryos of sea urchins Strongylocentrotus droebachiensis and S, intermedius}; Neifakh AA et al.; The time of expression of the genes controlling aldolase has been studied in the hybrid embryos female Strongylocentrotus droebachiensis X male S . intermedius . The enzyme heat resistance estimated by the temperature of 50% inactivation following the exposition for 30 min (T50) was used as its genetic marker . T50 of aldolase of the psychrophilic maternal species suffered practically no changes from the stage of mesenchyme blastula till the stage of 11 days old pluteus and equated 35.3 degrees . T50 of aldolase of autumn- and spring-spawning populations of the thermophilic paternal species equaled 39.5 degrees at the stage of 11 days old pluteus . The heat resistance of aldolase of the hybrid embryos did not differ reliably from that of maternal enzyme during the first 4 days of development (at 8 degrees) till the late gastrula stage and attained the maximum (T50 =36.9 degrees) on the 8th day (stage of pluteus) . The expression of the genes controlling aldolase appears to take place between these developmental stages. Prikl Biokhim Mikrobiol, 1976 Jan-Feb, 12(1), 24 - 9 {Peptidases of preparations of proteolytic enzymes from several species of bacteria, molds and actinomycetes}; Lysenkov NV; The activity of carboxy- and aminopeptidases of enzymic preparations obtained from bacteria Bacillus subtilis 1 M, 17, Bacillus mesentericus 100/26, 100/56, 16; Pseudomonas aeruginosa 1/7a, 1/4; Bacillus thermophilicus S . sp . nov B-22, moulds Aspergillus oryzae 1-746, Aspergillus flavus 716 and actinomycete Streptomyces griseus 32-13 was studied . In the bacterial preparations the proteolytic activity was 1.5-2.5 times lower than in the fungal and actinomycete preparations . The carboxypeptidase activity of bacterial preparations was comparable with that of the moulds and by one order of magnitude lower than that of the actinomycete . The aminopeptidase activity of the bacterial preparations was different; however, it was comparable with that of the moulds and was higher than that in some cases . In the Streptomyces preparation the aminopeptidase activity was significantly higher than in the bacterial and fungal preparations . In most preparations Co2+ and Mn2+ influenced the activity of carboxy- and amino-peptidases . This suggests that these proteins can be referred to metal enzymes. Experientia Suppl, 1976, 26, 77 - 89 Amylase activity and stability at high and low temperature depending on calcium and other divalent cations; Heinen W et al.; Since the extracellular amylase from B . caldolyticus had been characterized as a Ca-dependent enzyme, we wanted to determine the function and specificity of this ion, and it's possible significance for the thermophilic properties of the enzyme . The first question concerned the substitution of Ca with regard to the activity at 70 C . Our data show that Sr or other divalent cations can only to a limited extent replace Ca, and that only in presence of Ca full activity can be achieved . When the majority of the binding sites of the subunits are occupied by cations other than Ca, an almost total loss of activity results . The reason for this kind of inactivation was found in the responsibility of Ca for the stability of the enzyme at high temperature . The omission of Ca then leads to an irreversible denaturation of the enzyme, so that after this kind of treatment no activity is detectable at either high or low temperature, which means that the amylase is not a thermostable enzyme in the classical sense . The stabilizing effect of Ca could not be substituted by any other of the cations tested . Experiments concerning the stability of the enzyme at various temperatures in absence of presence of Ca revealed that the enzyme is stable up to 55 C as long as trace amounts of Ca are available . If these are omitted by a chelator, the enzyme becomes unstable between 40 and 45 C . Experiments in which a certain protein concentration range was tested at 40 and 70 C with given concentrations of Ca showed that the stability, and with it the activity, at high temperature is directly related to the amount of Ca available: The more the Ca supply is limited, the less enzyme protein can be kept in the right configuration, and irreversibly denatures as a result . At low temperature, however, the enzyme becomes almost independent of Ca, and the small amounts necessary to obtain full activity can be replaced by other divalent cations . The main conclusions concerning the role of Ca are that at high temperatures it participates in achieving the correct configuration of the thermostable form of the enzyme, and that in this function it is irreplacable . At low temperature it acts as an unspecific cofactor . The other conclusion is that the amylase can exist either as a caldo-active, thermostable, Ca-dependent enzyme, or in a Ca-independent thermostable state with a lower activity. Experientia Suppl, 1976, 26, 19 - 30 Thermal stability of homologous neutral metalloendopeptidases in thermophilic and mesophilic bacteria: structural considerations; Pangburn MK et al.; Thermolysin and neutral protease A are neutral metalloendopeptidases having similar specificity, molecular weight, metal content, and amino acid composition . Thermolysin, derived from the thermophilic organism Bacillus thermoproteolyticus, is heat inactivated at about 84 degrees whereas neutral protease A, derived from the mesophilic organism Bacillus subtilis, is inactivated at about 59 degrees . Structural analyses reveal that the two enzymes are homologous . Of the 326 residues of neutral protease A, 171 have been placed in sequence and 49% of these have been found in identical loci in thermolysin . These include many of the residues corresponding to the active site of thermolysin . The sensitivity of both enzymes to thermal inactivation is dependent upon the presence of calcium and neutral protease appears to bind less calcium than thermolysin . Structural data indicate that many of the ligands associated with calcium sites 1 and 2 (double site of thermolysin) are present in neutral protease and that calcium site 4 cannot exist in neutral protease . The structural homology and functional analogy of these two proteins support the concept that they have similar conformations . The known structure of thermolysin is used as a model to discuss structural differences which might be related to thermal stability. Biochimie, 1976, 58(1-2), 183 - 99 Purification and properties of two initiation factors from Bacillus stearothermophilus; Kay AC et al.; Two initiation factors have been isolated from the thermophilic bacterium, Bacillus stearothermophilus, and purified to near homogeneity . The two factors possess physical characteristics and activities associated with the E . coli initiation factors IF-2 and IF-3, and are interchangeable with these factors . The two systems present, however, several differences : S-IF-2 is significantly more heat stable than E . coli IF-2, loosing less than 50 per cent of its activity after 20 minutes at 70degreesC . S-IF-2 alone is unable to promote initiation complex formation on B . stearothermophilus or E . coli ribosomes, and S-IF-3 is absolutely necessary for initiation of complex formation on B . stearothermophilus ribosomes . No factor corresponding to IF-1 has been found . S-IF-3 appears to be able to replace at least partially IF-1, since S-IF-3 and E . coli IF-2 are sufficient to promote maximum fMet-tRNA binding to E . coli ribosomes, while E . coli IF-3 and IF-2 also require IF-1 . The differences between the two systems are perhaps required because of the elevated temperature at which B . stearothermophilus normally grows. Experientia Suppl, 1976, 26, 147 - 55 Comparative conformational properties of thermophilic and mesophilic 6-phosphogluconate dehydrogenase; Veronese FM et al.; The structural properties of 6-phosphogluconate dehydrogenase from the mesophilic bacterium E . coli and the thermophilic B . stearothermophilus are compared using circular dichroism and fluorescence emission spectroscopy . The enzymes appear to possess a similar structure which does not change on heating up to the respective temperature of stability of the enzyme . The thermostability of the two 6-phosphogluconate dehydrogenases as determined by activity measurements parallels that determined by CD with the melting profile method, indicating that the loss of biological activity in the enzymes is directly related to the unfolding of the protein molecule . The pattern of unfolding of the proteins by the action of 8 M urea suggests that a core of enhanced conformational stability exists in the B . stearothermophilus enzyme. Experientia Suppl, 1976, 26, 135 - 45 Thermal properties of glyceraldehyde 3-phosphate dehydrogenase from Escherichia coli; Fontana A et al.; The molecular properties of glyceraldehyde 3-phosphate dehydrogenase from E . coli have been evaluated by circular dichroism and fluorescence emission spectroscopy measurements, with the purpose of studying the structural properties which are relevant for a comparison with the enzyme from the obligate thermophile Bacillus stearothermophilus . The enzyme is moderately resistant to heat treatment, being pratically stable when treated for 10 min at 50 degrees C and completely inactivated when heating was performed at 60 degrees C . The secondary structure of the E . coli GPDH appears to be predominatly beta-structure as judged by circular dichroism, showing a negative band centered at about 219 nm . The emission fluorescence of the enzyme shows a maximum at 333 nm upon excitation at 295 nm . In the native E . coli enzyme the tryptophan residues seem to be buried in a hydrophobic region rather than exposed to a polar environment . The structure of the enzyme did not change up to about 50 degrees C, at which temperature thermal inactivation takes place . Upon denaturation the circular dichronic signal at 219 nm gradually decreases, and a red shift of the emission maximum from 333 nm to ca . 345 nm upon heating is indicative that the native structure of the enzyme is unfolded, the tryptophan being exposed to the solvent medium . Since it has been found that the E . coli GPDH closely resembles in many of its properties the B . stearothermophilus enzyme, this bacterial enzyme seems to be useful for comparision with the thermophilic enzyme in studies of its thermostability. Z Allg Mikrobiol, 1976, 19(5), 353 - 60 Properties of proteolytic enzymes isolated from a thermophilic strain of Micromonospora vulgaris 42; Nesterova NG et al.; Two proteolytic enzymes, protease A and protease B, were isolated in homogeneous state from the cultural broth of the thermophilic actinomycete Micromonospora vulgaris 42 . Their physicochemical properties were studied, i.e., molecular weight (50 000 for protease A and 30 000 for protease B), amino acid composition, N-terminal amino acids (phenylalanine for protease A and alanine for protease B) . The specificity of the action of these enzymes was assayed by splitting the B chain of oxidized insulin . Both enzymes are neutral proteases of the thermolysine type. Vet Med Nauki, 1976, 13(2), 39 - 48 {Development and resistance of staphylococci in Bulgaricus milk}; Dincheva E; Studied was the dynamics of development of 9 strains of staphilococci isolated from humans and animals, kept in heat-treated milk, and their resistance in Bulgarian sour milk . It was established that the pathogenic Staph . aureus and Staph . epidermidis develop will in fresh milk kept up to 7 days at 2--6degreesC and 18--22degreesC . In the production of Bulgarian sour milk Staph, aureus was shown to be viable, remaining active for seven days at 2 to 6degreesC . At room temperature (18--22degreesC) the survival rate has been dependent on the dynamics of accumulating metabolite products in connection with the development of Lactobac . bulgaricum and Streptococcus thermophilus . When the total acidity value reaches 160degreesT the pathogenic staphylococci are destroyed . Staphylococcus epidermidis finds no favourable medium to develop in Bulgarian sour milk, and it perishes when the total acidity is 120degreesT and pH -- 3. Experientia Suppl, 1976, 26, 187 - 97 Maintainance of specificity, information, and thermostability in thermophilic Bacillus sp . glutamine synthetase; Wedler FC et al.; Glutamine synthetase has been purified to homogeneity from B . subtilis (37 degrees) B . stearothermophilus (55 degrees), and B . caldolyticus (75 degrees) . Those characteristics compared include size (6.0 +/- 0.3 X 10(5) daltons), quaternary structure (12 SU) amino acid content, substrate Km's and specificity for structural analogs, metal ion activation, number and kind of separate feedback modifier sites, and the complexity of modifier-substrate and modifier-modifier site interactions . Although the 37 degrees and 55 degrees systems are quite similar, the 75 degrees system shows important alterations in substrate specificity and modes of modifier action . Whereas at 37 degrees and 55 degrees AMP inhibits synergistically with amino acids (glycine, glutamine, histidine), the 75 degrees enzyme is inhibited directly by the products ADP, (which assumes the role of AMP) and glutamine, plus other ligands . Ligand binding domains are compared and found to be very different . Thermostabilization occurs by (a) protection by bound L-glutamate, (b) protein aggregation, (c) trends in the content of total polar residues, total Asx + Flx residues, the average hydrophobicity, and (d) disulfide bond cross-linking . Such studies provide insights to molecular evolution occurring with changes in environmental stress. Experientia Suppl, 1976, 26, 169 - 83 Purification and some properties of NADP+ -specific isocitrate dehydrogenase from an extreme thermophile, Thermus flavus AT-62; Saiki T et al.; Thermostable NADP+ -specific isocitrate dehydrogenase (EC 1.1.1.42) was purified from crude extract of an extremely thermophilic bacterium Thermus flavus AT-62 through DEAE-cellulose column, acetone fractionation, DEAE-Sephadex A-50 column and isoelectric focussing . The enzyme was purified about 500-folds in its specific activity and purity was found to be about 96% . The enzyme was not inactivated after 60 min at 70 degrees C, but 20 and 80% of the activity were lost after 60 min at 80 degrees and 90 degrees C, respectively . Oxalacetate plus glyoxylate (each 1 nM) demonstrated 75% inhibition of the activity in concerted manner . The degree of the inhibition and the affinity of the enzyme for isocitrate and NADP+ decreased with the rise of temperature, especially above 60 degrees C . The activation energy below and above 60 degrees C were 14,500 and 8,000 cal per mole respectively . In CD spectra negative bands at 210 and 220nm were observed and alpha-helix content was calculated to be about 26% . In the course of heating up to 60 degrees practically no change in CD bands are observed, but above 60 degrees the depth of CD bands decreased gradually and remarkably above 80 degrees C . The effect of temperature on kinetic parameters and secondary structures of the enzyme was discussed in relation to the temperature adaptation of the organism.
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