|
|
|
|
|
|
Characterization of the 3-O-Methylgallate Dioxygenase Gene and Evidence of Multiple 3-O-Methylgallate Catabolic Pathways in Sphingomonas paucimobilis SYK-6. Daisuke Kasai, 2004.Sphingomonas paucimobilis SYK-6 is able to grow on various lignin-derived biaryls as the sole source of carbon and energy . These compounds are degraded to vanillate and syringate by the unique and specific enzymes in this strain . Vanillate and syringate are converted to protocatechuate (PCA) and 3-O-methylgallate (3MGA), respectively, by the tetrahydrofolate-dependent O-demethylases . Previous studies have suggested that these compounds are further degraded via the PCA 4,5-cleavage pathway . However, our subsequent analysis of the ligB insertion mutant, which encodes the ß subunit of PCA 4,5-dioxygenase, suggested that at least one alternative route is involved in 3MGA degradation . In the present study, we isolated the desZ gene, which confers 3MGA degradation activity on Escherichia coli . The deduced amino acid sequence of desZ showed ca . 20 to 43% identity with the type II extradiol dioxygenases . Gas chromatography-mass spectrometry analysis suggested that DesZ catalyzes the 3,4-cleavage of 3MGA . Disruption of both desZ and ligB in SYK-6 resulted in loss of the dioxygen-dependent 3MGA transformation activity, but the resulting mutant retained the ability to grow on syringate . We found that the cell extract of the desZ ligB double mutant was able to convert 3MGA to gallate when tetrahydrofolate was added to the reaction mixture, and the cell extract of this mutant degraded gallate to the same degree as the wild type did . All these results suggest that syringate is degraded through multiple 3MGA degradation pathways in which ligAB, desZ, 3MGA O-demethylase, and gallate dioxygenase are participants . The Evolving Genome of Salmonella enterica Serovar Pullorum. Gui-Rong Liu, 2002.Salmonella enterica serovar Pullorum is a fowl-adapted bacterial pathogen that causes dysentery (pullorum disease) . Host adaptation and special pathogenesis make S . enterica serovar Pullorum an exceptionally good system for studies of bacterial evolution and speciation, especially regarding pathogen-host interactions and the acquisition of pathogenicity . We constructed a genome map of S . enterica serovar Pullorum RKS5078, using I-CeuI, XbaI, AvrII, and SpeI and Tn10 insertions . Pulsed-field gel electrophoresis was employed to separate the large DNA fragments generated by the endonucleases . The genome is 4,930 kb, which is similar to most salmonellas . However, the genome of S . enterica serovar Pullorum RKS5078 is organized very differently from the majority of salmonellas, with three major inversions and one translocation . This extraordinary genome structure was seen in most S . enterica serovar Pullorum strains examined, with different structures in a minority of S . enterica serovar Pullorum strains . We describe the coexistence of different genome structures among the same bacteria as genomic plasticity . Through comparisons with S . enterica serovar Typhimurium, we resolved seven putative insertions and eight deletions ranging in size from 12 to 157 kb . The genomic plasticity seen among S . enterica serovar Pullorum strains supported our hypothesis about its association with bacterial evolution: a large genomic insertion (157 kb in this case) disrupted the genomic balance, and rebalancing by independent recombination events in individual lineages resulted in diverse genome structures . As far as the structural plasticity exists, the S . enterica serovar Pullorum genome will continue evolving to reach a further streamlined and balanced structure . Anaerobic Respiration Using Fe3+, S0, and H2 in the Chemolithoautotrophic Bacterium Acidithiobacillus ferrooxidans. Naoya Ohmura, 2002.The chemolithoautotrophic bacterium Acidithiobacillus ferrooxidans has been known as an aerobe that respires on iron and sulfur . Here we show that the bacterium could chemolithoautotrophically grow not only on H2/O2 under aerobic conditions but also on H2/Fe3+, H2/S0, or S0/Fe3+ under anaerobic conditions . Anaerobic respiration using Fe3+ or S0 as an electron acceptor and H2 or S0 as an electron donor serves as a primary energy source of the bacterium . Anaerobic respiration based on reduction of Fe3+ induced the bacterium to synthesize significant amounts of a c-type cytochrome that was purified as an acid-stable and soluble 28-kDa monomer . The purified cytochrome in the oxidized form was reduced in the presence of the crude extract, and the reduced cytochrome was reoxidized by Fe3+ . Respiration based on reduction of Fe3+ coupled to oxidation of a c-type cytochrome may be involved in the primary mechanism of energy production in the bacterium on anaerobic iron respiration . Development of Methicillin Resistance in Clinical Isolates of Staphylococcus sciuri by Transcriptional Activation of the mecA Homologue Native to the Species. Isabel Couto, 2003.The ß-lactam resistance gene mecA was acquired by Staphylococcus aureus from an extraspecies source . The search for the possible origin of this gene has led to the identification of a close structural homologue of mecA as a native gene in the animal species Staphylococcus sciuri. Surprisingly, the overwhelming majority of S . sciuri isolates were fully susceptible to ß-lactam antibiotics in spite of the ubiquitous presence of the mecA homologue in the bacteria . We now describe two unusual S . sciuri strains isolated from humans—SS-37 and SS-41—that showed resistance to methicillin associated with high rates of transcription of the mecA homologue and production of a protein resembling penicillin binding protein 2a, the gene product of S . aureus mecA . In strain SS-37 increased transcription of the mecA homologue was related to insertion of an IS256 element upstream of the structural gene, and strain SS-41 had single nucleotide alterations in the promoter region of the mecA homologue which appear to be related to up-regulation of the rate of transcription . A third methicillin-resistant human isolate of S . sciuri that carries both the native mecA homologue and a methicillin-resistant S . aureus (MRSA) type mecA, strain K3, was now shown to be unstable in the absence of drug selection, causing the segregation of antibiotic-susceptible cells accompanied by the loss of the MRSA type mecA . These observations illustrate the remarkable variety of strategies available to bacteria for acquiring mechanisms of drug resistance in the in vivo environment . Identification and Characterization of a Mandelamide Hydrolase and an NAD(P)+-Dependent Benzaldehyde Dehydrogenase from Pseudomonas putida ATCC 12633. Michael J. McLeish, 2003.The enzymes of the mandelate metabolic pathway permit Pseudomonas putida ATCC 12633 to utilize either or both enantiomers of mandelate as the sole carbon source . The genes encoding the mandelate pathway were found to lie on a single 10.5-kb restriction fragment . Part of that fragment was shown to contain the genes coding for mandelate racemase, mandelate dehydrogenase, and benzoylformate decarboxylase arranged in an operon . Here we report the sequencing of the remainder of the restriction fragment, which revealed three further open reading frames, denoted mdlX, mdlY, and mdlD . All were transcribed in the opposite direction from the genes of the mdlABC operon . Sequence alignments suggested that the open reading frames encoded a regulatory protein (mdlX), a member of the amidase signature family (mdlY), and an NAD(P)+-dependent dehydrogenase (mdlD) . The mdlY and mdlD genes were isolated and expressed in Escherichia coli, and the purified gene products were characterized as a mandelamide hydrolase and an NAD(P)+-dependent benzaldehyde dehydrogenase, respectively .
|
© 2005
Transgalactic Ltd (manufacturer of Bioscreen C software) |
Privacy Statement | P.O. Box
1393, 00101 Helsinki, Finland,
Last modified: May 25, 2005
| ||||||
