Advances in Microbiology最新文献

{"title":"Description of a Putative Oligosaccharyl:S-Layer Protein Transferase from the Tyrosine <i>O</i>-Glycosylation System of <i>Paenibacillus alvei</i> CCM 2051^T.","authors":"Robin Ristl,&nbsp;Bettina Janesch,&nbsp;Julia Anzengruber,&nbsp;Agnes Forsthuber,&nbsp;Johanna Blaha,&nbsp;Paul Messner,&nbsp;Christina Schäffer","doi":"10.4236/aim.2012.24069","DOIUrl":"https://doi.org/10.4236/aim.2012.24069","url":null,"abstract":"<p><p>Surface (S)-layer proteins are model systems for studying protein glycosylation in bacteria and simultaneously hold promises for the design of novel, glyco-functionalized modules for nanobiotechnology due to their 2D self-assembly capability. Understanding the mechanism governing S-layer glycan biosynthesis in the Gram-positive bacterium <i>Paenibacillus alvei</i> CCM 2051^T is necessary for the tailored glyco-functionalization of its S-layer. Here, the putative oligosaccharyl:S-layer protein transferase WsfB from the <i>P. alvei</i> S-layer glycosylation gene locus is characterized. The enzyme is proposed to catalyze the final step of the glycosylation pathway, transferring the elongated S-layer glycan onto distinct tyrosine <i>O</i>-glycosylation sites. Genetic knock-out of WsfB is shown to abolish glycosylation of the S-layer protein SpaA but not that of other glycoproteins present in <i>P. alvei</i> CCM 2051^T, confining its role to the S-layer glycosylation pathway. A transmembrane topology model of the 781-amino acid WsfB protein is inferred from activity measurements of green fluorescent protein and phosphatase A fused to defined truncations of WsfB. This model shows an overall number of 13 membrane spanning helices with the Wzy_C domain characteristic of <i>O</i>-oligosaccharyl:protein transferases (<i>O</i>-OTases) located in a central extra-cytoplasmic loop, which both compares well to the topology of OTases from Gram-negative bacteria. Mutations in the Wzy_C motif resulted in loss of WsfB function evidenced in reconstitution experiments in <i>P. alvei</i> ΔWsfB cells. Attempts to use WsfB for transferring heterologous oligosaccharides to its native S-layer target protein in <i>Escherichia coli</i> CWG702 and <i>Salmonella enterica</i> SL3749, which should provide lipid-linked oligosaccharide substrates mimicking to some extent those of the natural host, were not successful, possibly due to the stringent function of WsfB. Concluding, WsfB has all features of a bacterial <i>O</i>-OTase, making it the most probable candidate for the oligosaccharyl:S-layer protein transferase of <i>P. alvei</i>, and a promising candidate for the first <i>O</i>-OTase reported in Gram-positives.</p>","PeriodicalId":7355,"journal":{"name":"Advances in Microbiology","volume":"2 4","pages":"537-546"},"PeriodicalIF":0.0,"publicationDate":"2012-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4397953/pdf/emss-62937.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"33115048","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Differential Role of Two-Component Regulatory Systems (<i>phoPQ</i> and <i>pmrAB</i>) in Polymyxin B Susceptibility of <i>Pseudomonas aeruginosa.</i>","authors":"Daniel Owusu-Anim,&nbsp;Dong H Kwon","doi":"10.4236/aim.2012.21005","DOIUrl":"https://doi.org/10.4236/aim.2012.21005","url":null,"abstract":"<p><p>Polymyxins are often considered as a last resort to treat multidrug resistant <i>P. aeruginosa</i> but polymyxin resistance has been increasingly reported worldwide in clinical isolates. Polymyxin resistance in <i>P. aeruginosa</i> is known to be associated with alterations in either PhoQ or PmrB. In this study, mutant strains of <i>P. aeruginosa</i> carrying amino acid substitution, a single and/or dual inactivation of PhoQ and PmrB were constructed to further understand the roles of PhoQ and PmrB in polymyxin susceptibility. Polymyxin B resistance was caused by both inactivation and/or amino acid substitutions in PhoQ but by only amino acid substitutions of PmrB. Alterations of both PhoQ and PmrB resulted in higher levels of polymyxin B resistance than alteration of either PhoQ or PmrB alone. These results were confirmed by time-killing assays suggesting that high-level polymyxin resistance in <i>P. aeruginosa</i> is caused by alterations of both PhoQ and PmrB.</p>","PeriodicalId":7355,"journal":{"name":"Advances in Microbiology","volume":"2 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2012-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3859615/pdf/nihms528316.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"31965972","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}