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Cold Shock Response in Sporulating Bacillus subtilis and Its Effect on Spore Heat Resistance. Sara Movahedi, 2002.Cold shock and ethanol and puromycin stress responses in sporulating Bacillus subtilis cells have been investigated . We show that a total of 13 proteins are strongly induced after a short cold shock treatment of sporulating cells . The cold shock pretreatment affected the heat resistance of the spores formed subsequently, with spores heat killed at 85 or 90°C being more heat resistant than the control spores while they were more heat sensitive than controls that were heat treated at 95 or 100°C . However, B . subtilis spores with mutations in the main cold shock proteins, CspB, -C, and -D, did not display decreased heat resistance compared to controls, indicating that these proteins are not directly responsible for the increased heat resistance of the spores . The disappearance of the stress proteins later in sporulation suggests that they cannot be involved in repairing heat damage during spore germination and outgrowth but must alter spore structure in a way which increases or decreases heat resistance . Since heat, ethanol, and puromycin stress produce similar proteins and similar changes in spore heat resistance while cold shock is different in both respects, these alterations appear to be very specific . raiIR Genes Are Part of a Quorum-Sensing Network Controlled by cinI and cinR in Rhizobium leguminosarum. F. Wisniewski-Dyé, 2002.Analysis of N-acyl-L-homoserine lactones (AHLs) produced by Rhizobium leguminosarum bv . viciae indicated that there may be a network of quorum-sensing regulatory systems producing multiple AHLs in this species . Using a strain lacking a symbiosis plasmid, which carries some of the quorum-sensing genes, we isolated mutations in two genes (raiI and raiR) that are required for production of AHLs . The raiIR genes are located adjacent to dad genes (involved in D-alanine catabolism) on a large indigenous plasmid . RaiR is predicted to be a typical LuxR-type quorum-sensing regulator and is required for raiI expression . The raiR gene was expressed at a low level, possibly from a constitutive promoter, and its expression was increased under the influence of the upstream raiI promoter . Using gene fusions and analysis of AHLs produced, we showed that expression of raiI is strongly reduced in strains carrying mutations in cinI or cinR, genes which determine a higher-level quorum-sensing system that is required for normal expression of raiIR . The product of CinI, N-(3-hydroxy-7-cis tetradecenoyl) homoserine lactone, can induce raiR-dependent raiI expression, although higher levels of expression are induced by other AHLs . Expression of raiI in a strain of Agrobacterium that makes no AHLs resulted in the identification of N-(3-hydroxyoctanoyl)-L-homoserine lactone (3OH,C8-HSL) as the major product of RaiI, although other AHLs that comigrate with N-hexanoyl-, N-heptanoyl-, and N-octanoyl-homoserine lactones were also made at low levels . The raiI gene was strongly induced by 3OH,C8-HSL (the product of RaiI) but could also be induced by other AHLs, suggesting that the raiI promoter can be activated by other quorum-sensing systems within a network of regulation which also involves AHLs determined by genes on the symbiotic plasmid . Thus, the raiIR and cinIR genes are part of a complex regulatory network that influences AHL biosynthesis in R . leguminosarum . Lactobacillus reuteri CRL1098 Produces Cobalamin. María P. Taranto, 2003.We found that Lactobacillus reuteri CRL1098, a lactic acid bacterium isolated from sourdough, is able to produce cobalamin . The sugar-glycerol cofermentation in vitamin B12-free medium showed that this strain was able to reduce glycerol through a well-known cobalamin-dependent reaction with the formation of 1,3-propanediol as a final product . The cell extract of L . reuteri corrected the coenzyme B12 requirement of Lactobacillus delbrueckii subsp . lactis ATCC 7830 and allowed the growth of Salmonella enterica serovar Typhimurium (metE cbiB) and Escherichia coli (metE) in minimal medium . Preliminary genetic studies of cobalamin biosynthesis genes from L . reuteri allowed the identification of cob genes which encode the CobA, CbiJ, and CbiK enzymes involved in the cobalamin pathway . The cobamide produced by L . reuteri, isolated in its cyanide form by using reverse-phase high-pressure liquid chromatography, showed a UV-visible spectrum identical to that of standard cyanocobalamin (vitamin B12) .
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