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The Enterococcus faecalis fsr Two-Component System Controls Biofilm Development through Production of Gelatinase. Lynn E. Hancock, 2004.Bacterial growth as a biofilm on solid surfaces is strongly associated with the development of human infections . Biofilms on native heart valves (infective endocarditis) is a life-threatening disease as a consequence of bacterial resistance to antimicrobials in such a state . Enterococci have emerged as a cause of endocarditis and nosocomial infections despite being normal commensals of the gastrointestinal and female genital tracts . We examined the role of two-component signal transduction systems in biofilm formation by the Enterococcus faecalis V583 clinical isolate and identified the fsr regulatory locus as the sole two-component system affecting this unique mode of bacterial growth . Insertion mutations in the fsr operon affected biofilm formation on two distinct abiotic surfaces . Inactivation of the fsr-controlled gene gelE encoding the zinc-metalloprotease gelatinase was found to prevent biofilm formation, suggesting that this enzyme may present a unique target for therapeutic intervention in enterococcal endocarditis . Origins of the 2,4-Dinitrotoluene Pathway. Glenn R. Johnson, 2002.The degradation of synthetic compounds requires bacteria to recruit and adapt enzymes from pathways for naturally occurring compounds . Previous work defined the steps in 2,4-dinitrotoluene (2,4-DNT) metabolism through the ring fission reaction . The results presented here characterize subsequent steps in the pathway that yield the central metabolic intermediates pyruvate and propionyl coenzyme A (CoA) . The genes encoding the degradative pathway were identified within a 27-kb region of DNA cloned from Burkholderia cepacia R34, a strain that grows using 2,4-DNT as a sole carbon, energy, and nitrogen source . Genes for the lower pathway in 2,4-DNT degradation were found downstream from dntD, the gene encoding the extradiol ring fission enzyme of the pathway . The region includes genes encoding a CoA-dependent methylmalonate semialdehyde dehydrogenase (dntE), a putative NADH-dependent dehydrogenase (ORF13), and a bifunctional isomerase/hydrolase (dntG) . Results from analysis of the gene sequence, reverse transcriptase PCR, and enzyme assays indicated that dntD dntE ORF13 dntG composes an operon that encodes the lower pathway . Additional genes that were uncovered encode the 2,4-DNT dioxygenase (dntAaAbAcAd), methylnitrocatechol monooxygenase (dntB), a putative LysR-type transcriptional (ORF12) regulator, an intradiol ring cleavage enzyme (ORF3), a maleylacetate reductase (ORF10), a complete ABC transport complex (ORF5 to ORF8), a putative methyl-accepting chemoreceptor protein (ORF11), and remnants from two transposable elements . Some of the additional gene products might play as-yet-undefined roles in 2,4-DNT degradation; others appear to remain from recruitment of the neighboring genes . The presence of the transposon remnants and vestigial genes suggests that the pathway for 2,4-DNT degradation evolved relatively recently because the extraneous elements have not been eliminated from the region . DNA Array-Based Transcriptional Analysis of Asporogenous, Nonsolventogenic Clostridium acetobutylicum Strains SKO1 and M5. Christopher A. Tomas, 2003.The large-scale transcriptional program of two Clostridium acetobutylicum strains (SKO1 and M5) relative to that of the parent strain (wild type [WT]) was examined by using DNA microarrays . Glass DNA arrays containing a selected set of 1,019 genes (including all 178 pSOL1 genes) covering more than 25% of the whole genome were designed, constructed, and validated for data reliability . Strain SKO1, with an inactivated spo0A gene, displays an asporogenous, filamentous, and largely deficient solventogenic phenotype . SKO1 displays downregulation of all solvent formation genes, sigF, and carbohydrate metabolism genes (similar to genes expressed as part of the stationary-phase response in Bacillus subtilis) but also several electron transport genes . A major cluster of genes upregulated in SKO1 includes abrB, the genes from the major chemotaxis and motility operons, and glycosylation genes . Strain M5 displays an asporogenous and nonsolventogenic phenotype due to loss of the megaplasmid pSOL1, which contains all genes necessary for solvent formation . Therefore, M5 displays downregulation of all pSOL1 genes expressed in the WT . Notable among other genes expressed more highly in WT than in M5 were sigF, several two-component histidine kinases, spo0A, cheA, cheC, many stress response genes, fts family genes, DNA topoisomerase genes, and central-carbon metabolism genes . Genes expressed more highly in M5 include electron transport genes (but different from those downregulated in SKO1) and several motility and chemotaxis genes . Most of these expression patterns were consistent with phenotypic characteristics . Several of these expression patterns are new or different from what is known in B . subtilis and can be used to test a number of functional-genomic hypotheses . Development of a mariner-Based Transposon for Use in Sorangium cellulosum. Bryan Julien, 2003.In order to generate marked insertions in the myxobacterium Sorangium cellulosum, a transposon based on the eukaryotic mariner transposon was developed . The transposition frequency was increased with the use of a mutated tnp gene . The transposon randomly inserts into the chromosome, as demonstrated by targeted mutagenesis of the epoK gene .
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