|
|
|
|
|
|
pH-Dependent Catabolic Protein Expression during Anaerobic Growth of Escherichia coli K-12. Elizabeth Yohannes, 2004.During aerobic growth of Escherichia coli, expression of catabolic enzymes and envelope and periplasmic proteins is regulated by pH . Additional modes of pH regulation were revealed under anaerobiosis . E . coli K-12 strain W3110 was cultured anaerobically in broth medium buffered at pH 5.5 or 8.5 for protein identification on proteomic two-dimensional gels . A total of 32 proteins from anaerobic cultures show pH-dependent expression, and only four of these proteins (DsbA, TnaA, GatY, and HdeA) showed pH regulation in aerated cultures . The levels of 19 proteins were elevated at the high pH; these proteins included metabolic enzymes (DhaKLM, GapA, TnaA, HisC, and HisD), periplasmic proteins (ProX, OppA, DegQ, MalB, and MglB), and stress proteins (DsbA, Tig, and UspA) . High-pH induction of the glycolytic enzymes DhaKLM and GapA suggested that there was increased fermentation to acids, which helped neutralize alkalinity . Reporter lac fusion constructs showed base induction of sdaA encoding serine deaminase under anaerobiosis; in addition, the glutamate decarboxylase genes gadA and gadB were induced at the high pH anaerobically but not with aeration . This result is consistent with the hypothesis that there is a connection between the gad system and GabT metabolism of 4-aminobutanoate . On the other hand, 13 other proteins were induced by acid; these proteins included metabolic enzymes (GatY and AckA), periplasmic proteins (TolC, HdeA, and OmpA), and redox enzymes (GuaB, HmpA, and Lpd) . The acid induction of NikA (nickel transporter) is of interest because E . coli requires nickel for anaerobic fermentation . The position of the NikA spot coincided with the position of a small unidentified spot whose induction in aerobic cultures was reported previously; thus, NikA appeared to be induced slightly by acid during aeration but showed stronger induction under anaerobic conditions . Overall, anaerobic growth revealed several more pH-regulated proteins; in particular, anaerobiosis enabled induction of several additional catabolic enzymes and sugar transporters at the high pH, at which production of fermentation acids may be advantageous for the cell . Novel Type V Staphylococcal Cassette Chromosome mec Driven by a Novel Cassette Chromosome Recombinase, ccrC. Teruyo Ito, 2004.Staphylococcal cassette chromosome mec (SCCmec) is a mobile genetic element composed of the mec gene complex, which encodes methicillin resistance, and the ccr gene complex, which encodes the recombinases responsible for its mobility . The mec gene complex has been classified into four classes, and the ccr gene complex has been classified into three allotypes . Different combinations of mec gene complex classes and ccr gene complex types have so far defined four types of SCCmec elements . Now we introduce the fifth allotype of SCCmec, which was found on the chromosome of a community-acquired methicillin-resistant Staphylococcus aureus strain (strain WIS [WBG8318]) isolated in Australia . The element shared the same chromosomal integration site with the four extant types of SCCmec and the characteristic nucleotide sequences at the chromosome-SCCmec junction regions . The novel SCCmec carried mecA bracketed by IS431 (IS431-mecA-  mecR1-IS431), which is designated the class C2 mec gene complex; and instead of ccrA and ccrB genes, it carried a single copy of a gene homologue that encoded cassette chromosome recombinase . Since the open reading frame (ORF) was found to encode an enzyme which catalyzes the precise excision as well as site- and orientation-specific integration of the element, we designated the ORF cassette chromosome recombinase C (ccrC), and we designated the element type V SCCmec . Type V SCCmec is a small SCCmec element (28 kb) and does not carry any antibiotic resistance genes besides mecA . Unlike the extant SCCmec types, it carries a set of foreign genes encoding a restriction-modification system that might play a role in the stabilization of the element on the chromosome .
Analysis of the Peptidoglycan Hydrolase Complement of Lactococcus lactis: Identification of a Third N-Acetylglucosaminidase, AcmC. Carine Huard, 2004.The peptidoglycan hydrolase (PGH) complement of Lactococcus lactis was identified by amino acid sequence similarity searching of the L . lactis IL-1403 complete genome sequence . Five PGHs that are not encoded by prophages were detected, including the previously characterized AcmA and AcmB proteins . Four of these PGHs, AcmA to AcmD, contain a catalytic domain homologous to that of enterococcal muramidase, but they have different domain structures . The fifth one (YjgB) has sequence similarity with the active-site domain of peptidoglycan-specific endopeptidases . The three new PGH-encoding genes identified in this study are all actively transcribed in L . lactis subsp . cremoris MG1363 . The relative abundance of their transcripts varied during growth and was maximal during the early exponential growth phase . The three encoded proteins have peptidoglycan-hydrolyzing activities which are detected only at acidic pHs by zymography . Like AcmA and AcmB, AcmC has N-acetylglucosaminidase activity rather than the N-acetylmuramidase activity predicted by sequence similarity . Targeting of DMinC/MinD and DMinC/DicB Complexes to Septal Rings in Escherichia coli Suggests a Multistep Mechanism for MinC-Mediated Destruction of Nascent FtsZ Rings. Jay E. Johnson, 2002.The MinC protein is an important determinant of septal ring positioning in Escherichia coli . The N-terminal domain (ZMinC) suppresses septal ring formation by interfering with FtsZ polymerization, whereas the C-terminal domain (DMinC) is required for dimerization as well as for interaction with the MinD protein . MinD oscillates between the membrane of both cell halves in a MinE-dependent fashion . MinC oscillates along with MinD such that the time-integrated concentration of ZMinC at the membrane is minimal, and hence the stability of FtsZ polymers is maximal, at the cell center . MinC is cytoplasmic and fails to block FtsZ assembly in the absence of MinD, indicating that recruitment of MinC by MinD to the membrane enhances ZMinC function . Here, we present evidence that the binding of DMinC to MinD endows the MinC/MinD complex with a more specific affinity for a septal ring-associated target in vivo . Thus, MinD does not merely attract MinC to the membrane but also aids MinC in specifically binding to, or in close proximity to, the substrate of its ZMinC domain . MinC-mediated division inhibition can also be activated in a MinD-independent fashion by the DicB protein of cryptic prophage Kim . DicB shows little homology to MinD, and how it stimulates MinC function has been unclear . Similar to the results obtained with MinD, we find that DicB interacts directly with DMinC, that the DMinC/DicB complex has a high affinity for some septal ring target(s), and that MinC/DicB interferes with the assembly and/or integrity of FtsZ rings in vivo . The results suggest a multistep mechanism for the activation of MinC-mediated division inhibition by either MinD or DicB and further expand the number of properties that can be ascribed to the Min proteins . 2-Oxoglutarate:NADP+ Oxidoreductase in Azoarcus evansii: Properties and Function in Electron Transfer Reactions in Aromatic Ring Reduction. Christa Ebenau-Jehle, 2003.The conversion of [14C]benzoyl-coenzyme A (CoA) to nonaromatic products in the denitrifying ß-proteobacterium Azoarcus evansii grown anaerobically on benzoate was investigated . With cell extracts and 2-oxoglutarate as the electron donor, benzoyl-CoA reduction occurred at a rate of 10 to 15 nmol min-1 mg-1 . 2-Oxoglutarate could be replaced by dithionite (200% rate) and by NADPH (  10% rate); in contrast NADH did not serve as an electron donor . Anaerobic growth on aromatic compounds induced 2-oxoglutarate:acceptor oxidoreductase (KGOR), which specifically reduced NADP+, and NADPH:acceptor oxidoreductase . KGOR was purified by a 76-fold enrichment . The enzyme had a molecular mass of 290 ± 20 kDa and was composed of three subunits of 63 ( ), 62 ( ), and 37 (ß) kDa in a 1:1:1 ratio, suggesting an (ß)2 composition . The native enzyme contained Fe (24 mol/mol of enzyme), S (23 mol/mol), flavin adenine dinucleotide (FAD; 1.4 mol/mol), and thiamine diphosphate (0.95 mol/mol) . KGOR from A . evansii was highly specific for 2-oxoglutarate as the electron donor and accepted both NADP+ and oxidized viologens as electron acceptors; in contrast NAD+ was not reduced . These results suggest that benzoyl-CoA reduction is coupled to the complete oxidation of the intermediate acetyl-CoA in the tricarboxylic acid cycle . Electrons generated by KGOR can be transferred to both oxidized ferredoxin and NADP+, depending on the cellular needs . N-terminal amino acid sequence analysis revealed that the open reading frames for the three subunits of KGOR are similar to three adjacently located open reading frames in Bradyrhizobium japonicum . We suggest that these genes code for a very similar three-subunit KGOR, which may play a role in nitrogen fixation . The
-subunit is supposed to harbor one FAD molecule, two [4Fe-4S] clusters, and the NADPH binding site; the ß-subunit is supposed to harbor one thiamine diphosphate molecule and one further [4Fe-4S] cluster; and the
-subunit is supposed to harbor the CoA binding site . This is the first study of an NADP+-specific KGOR . A similar NADP+-specific pyruvate oxidoreductase, which contains all domains in one large subunit, has been reported for the mitochondrion of the protist Euglena gracilis and the apicomplexan Cryptosporidium parvum .
Microbial Isotopic Fractionation of Perchlorate Chlorine. Max L. Coleman, 2003.Perchlorate contamination can be microbially respired to innocuous chloride and thus can be treated effectively . However, monitoring a bioremediative strategy is often difficult due to the complexities of environmental samples . Here we demonstrate that microbial respiration of perchlorate results in a significant fractionation (  -15 ) of the chlorine stable isotope composition of perchlorate . This can be used to quantify the extent of biotic degradation and to separate biotic from abiotic attenuation of this contaminant .
|
© 2005
Transgalactic Ltd (manufacturer of Bioscreen C software) |
Privacy Statement | P.O. Box
1393, 00101 Helsinki, Finland,
Last modified: May 25, 2005
| ||||||
