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Transcriptional Regulation of Genes Encoding Arabinan-Degrading Enzymes in Bacillus subtilis. Maria Paiva Raposo, 2004.Bacillus subtilis produces hemicellulases capable of releasing arabinosyl oligomers and arabinose from plant cell walls . Inthis work, we characterize the transcriptional regulation ofthree genes encoding arabinan-degrading enzymes that are clusteredwith genes encoding enzymes that further catabolize arabinose.The abfA gene comprised in the metabolic operon araABDLMNPQ-abfA and the xsa gene located 23 kb downstream most probably encode alpha-L-arabinofuranosidases [EC 3.2.1.55] . Here, we show that theabnA gene, positioned immediately upstream from the metabolicoperon, encodes an endo-alpha-1,5-arabinanase [EC 3.2.1.99] . Furthermore,by in vivo RNA studies, we inferred that abnA and xsa are monocistronicand are transcribed from sigmaA-like promoters . Transcriptional fusionanalysis revealed that the expression of the three arabinasesis induced by arabinose and arabinan and is repressed by glucose.The levels of induction by arabinose and arabinan are higherduring early postexponential growth, suggesting a temporal regulation.Moreover, the induction mechanism of these genes is mediatedthrough negative control by the key regulator of arabinose metabolism,AraR . Thus, we analyzed AraR-DNA interactions by in vitro quantitativeDNase I footprinting and in vivo analysis of single-base-pairsubstitutions within the promoter regions of xsa and abnA . Theresults indicate that transcriptional repression of the abfAand xsa genes is achieved by a tightly controlled mechanismbut that the regulation of abnA is more flexible . We suggestthat the expression of genes encoding extracellular degradingenzymes of arabinose-containing polysaccharides, transport systems,and intracellular enzymes involved in further catabolism isregulated by a coordinate mechanism triggered by arabinose viaAraR. Regulation of Salmonella enterica Serovar Typhimurium mntH Transcription by H2O2, Fe2+, and Mn2+. David G. Kehres, 2002.MntH, a bacterial homolog of mammalian natural resistance associated macrophage protein 1 (Nramp1), is a primary transporter for Mn2+ influx in Salmonella enterica serovar Typhimurium and Escherichia coli . S . enterica serovar Typhimurium MntH contributes to H2O2 resistance and is important for full virulence . Consistent with its phenotype and function, mntH is regulated at the transcriptional level by both H2O2 and substrate cation . We have now identified three trans-acting regulatory factors and the three corresponding cis-acting mntH promoter motifs that mediate this regulation . In the presence of hydrogen peroxide, mntH is activated by OxyR, acting through an OxyR-binding motif centered just upstream of the likely -35 RNA polymerase-binding site . In the presence of Fe2+, mntH is repressed primarily by Fur, acting through a Fur-binding motif overlapping the -35 region . In the presence of Mn2+, mntH is repressed primarily by the Salmonella equivalent of E . coli b0817, a distant homolog of the Bacillus subtilis manganese transport repressor, MntR, acting through an inverted-repeat motif located between the likely -10 polymerase binding site and the ribosome binding site . E . coli b0817 was recently shown to bind the identical inverted-repeat motif in the E . coli mntH promoter and hence has been renamed MntR (S . I . Patzer and K . Hantke, J . Bacteriol . 183:4806-4813, 2001) . Using  fur,
mntR, and
fur
mntR mutant strains as well as mutations in the Fur- and MntR-binding motif elements, we found that Fe2+ can also mediate repression through the Mn2+ repressor MntR .
Evidence for a Type III Secretion System in Aeromonas salmonicida subsp . salmonicida. Sarah E. Burr, 2002. Functional Dissection of the XpsN (GspC) Protein of the Xanthomonas campestris pv . campestris Type II Secretion Machinery. Hsien-Min Lee, 2004.Type II secretion machinery is composed of 12 to 15 proteins for translocating extracellular proteins across the outer membrane . XpsL, XpsM, and XpsN are components of such machinery in the plant pathogen Xanthomonas campestris pv . campestris . All are bitopic cytoplasmic-membrane proteins, each with a large C-terminal periplasmic domain . They have been demonstrated to form a dissociable ternary complex . By analyzing the C-terminally truncated XpsN and PhoA fusions, we discovered that truncation of the C-terminal 103 residues produced a functional protein, albeit present below detectable levels . Furthermore, just the first 46 residues, encompassing the membrane-spanning sequence (residues 10 to 32), are sufficient to keep XpsL and XpsM at normal abundance . XpsN46(His6), synthesized in Escherichia coli, is able to associate in a membrane-mixing experiment with the XpsL-XpsM complex preassembled in X . campestris pv . campestris . The XpsN N-terminal 46 residues are apparently sufficient not only for maintaining XpsL and XpsM at normal levels but also for their stable association . The membrane-spanning sequence of XpsN was not replaceable by that of TetA . However, coimmunoprecipitation with XpsL and XpsM was observed for XpsN97::PhoA, but not XpsN46::PhoA . Only XpsN97::PhoA is dominant negative . Single alanine substitutions for three charged residues within the region between residues 47 and 97 made the protein nonfunctional . In addition, the R78A mutant XpsN protein was pulled down by XpsL-XpsM(His6) immobilized on an Ni-nitrilotriacetic acid column to a lesser extent than the wild-type XpsN . Therefore, in addition to the N-terminal 46 residues, the region between residues 47 and 97 of XpsN probably also plays an important role in interaction with XpsL-XpsM .
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