In addition to its role as carbon and energy source, fructose metabolism was reported to affect other cellular processes, such as biofilm formation by streptococci and bacterial pathogenicity in plants. Fructose genes encoding a 1-phosphofructokinase and a phosphotransferase system (PTS) fructose-specific enzyme IIABC component reside commonly in a gene cluster with a DeoR family regulator in various gram-positive bacteria. We present a comprehensive study of fructose metabolism in Lactococcus lactis, including a systematic study of fru mutants, global messenger analysis, and a molecular characterization of its regulation. The fru operon is regulated at the transcriptional level by both FruR and CcpA and at the metabolic level by inducer exclusion. The FruR effector is fructose-1-phosphate (F1P), as shown by combined analysis of transcription and measurements of the intracellular F1P pools in mutants either unable to produce this metabolite or accumulating it. The regulation of the fru operon by FruR requires four adjacent 10-bp direct repeats. The well-conserved organization of the fru promoter region in various low-GC gram-positive bacteria, including CRE boxes as well as the newly defined FruR motif, suggests that the regulation scheme defined in L. lactis could be applied to these bacteria. Transcriptome profiling of fruR and fruC mutants revealed that the effect of F1P and FruR regulation is limited to the fru operon in L. lactis. This result is enforced by the fact that no other targets for FruR were found in the available low-GC gram-positive bacteria genomes, suggesting that additional phenotypical effects due to fructose metabolism do not rely directly on FruR control, but rather on metabolism.Carbohydrate utilization systems are of particular importance to provide carbon and energy to bacteria. Among them, glucose and lactose systems have been widely studied, while other sugar utilization systems have been studied mostly with regard to their implication in targeted processes. Considering fructose availability in most ecosystems associated with plants, fructose metabolism and its regulation received little attention to date. The utilization of fructose is best documented in Escherichia coli, with the existence of three routes. In the main route, fructose is taken up via the membrane-spanning protein FruA and concomitantly phosphorylated to fructose-1-phosphate. Phosphorylation takes place by transfer of the phosphate group from phosphoenolpyruvate to fructose, which involves concerted action of two cytoplasmic proteins, EI of the phosphotransferase system (PTS) and a membrane-associated diphosphoryl transfer protein (FruB). The fructose-1-phosphate thus formed is further phosphorylated by ATP and 1-phosphofructokinase (FruK) to fructose-1,6-bisphosphate (17). The fruBKA operon of enteric bacteria is regulated at the transcriptional level primarily by the catabolite repressor-activator Cra (previously designated FruR) and by the cyclic AMP-CRP complex, which plays a secondary role (9,25).Although non-PTS...

No abstract

SummaryThe flagellar machinery is a highly complex organelle composed of a free rotating flagellum and a fixed stator that converts energy into movement. The assembly of the flagella and the stator requires interactions with the peptidoglycan layer through which the organelle has to pass for externalization. Lytic transglycosylases are peptidoglycan degrading enzymes that cleave the sugar backbone of peptidoglycan layer. We show that an endogenous lytic transglycosylase is required for full motility of Helicobacter pylori and colonization of the gastric mucosa. Deficiency of motility resulted from a paralysed phenotype implying an altered ability to generate flagellar rotation. Similarly, another Gram-negative pathogen Salmonella typhimurium and the Gram-positive pathogen Listeria monocytogenes required the activity of lytic transglycosylases, Slt or MltC, and a glucosaminidase (Auto), respectively, for full motility. Furthermore, we show that in absence of the appropriate lytic transglycosylase, the flagellar motor protein MotB from H. pylori does not localize properly to the bacterial pole. We present a new model involving the maturation of the surrounding peptidoglycan for the proper anchoring and functionality of the flagellar motor.

Aims: Staphylococcus carnosus, used as starter culture in fermented meat products, decreases the level of volatiles arising from lipid oxidation. To analyse its antioxidant capacities, catalase and superoxide dismutase (SOD) were characterized. Methods and Results: Catalase and SOD activities were measured with spectrophotometric methods and visualized on non-denaturing polyacrylamide gels. The corresponding sod gene was identi®ed by PCR. Southern hybridizations and enzymatic analyses showed that there was a single catalase and a single SOD in Staph. carnosus 833 strain. The gene encoding the Staph. carnosus SOD was found to encode a protein closely related to SOD requiring manganese. Catalase and SOD levels increased in mid-log phase. Only catalase was induced by oxygen, nitrate or nitrite while glucose induced neither enzyme. Metal ion limitation increased catalase and decreased SOD activities. Conclusions: Staph. carnosus synthesizes both enzymes in conditions encountered in sausage manufacturing. These results could explain the antioxidant properties of Staph. carnosus starter culture. Signi®cance and Impact of the Study: The knowledge of the antioxidant properties of Staphylococci will allow a more rational use of these starters in meat fermented products.

Staphylococcus xylosus is a facultative anaerobic bacterium used as a starter culture for fermented meat products. In an attempt to analyze the antioxidant capacities of this organism, the superoxide dismutase (SOD) was characterized. S. xylosus contains a single cytoplasmic SOD, which was not inhibited by H 2 O 2 . The SOD activity in crude extracts was completely lost upon metal depletion, but it could be recovered by manganese and very weakly by iron. It is therefore suggested that the S. xylosus SOD is a manganese-preferring enzyme. The corresponding gene, sod, was isolated from a genomic library of S. xylosus DNA and complemented the growth defect of an Escherichia coli SOD-deficient mutant. As deduced from the nucleotide sequence, sod encodes a protein of 199 amino acids with a molecular mass of 22.5 kDa. Two transcriptional start sites 25 and 120 bp upstream of the sod start codon were identified. A terminator-like structure downstream of the gene suggested a monocistronic sod mRNA. Regulation of sod expression was studied using fusions of the sod promoters to a genomic promoterless ␤-galactosidase gene. The sod expression was not affected by manganese and increased slightly with paraquat. It was induced during stationary phase in a complex medium but not in a chemically defined medium. To investigate the physiological role of SOD, a mutant devoid of SOD activity was constructed. Growth experiments showed that sod is not essential for aerobic growth in complex medium. However, in chemically defined medium without leucine, isoleucine, and valine, the sod mutant hardly grew, in contrast to the wild-type strain. In addition, the mutant was sensitive to hyperbaric oxygen and to paraquat. Therefore, sod plays an important role in the protection of S. xylosus from oxidative stress.