Wilson Marlena M, Anderson D Eric, Bernstein Harris D
Genetics and Biochemistry Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, United States of America.
Advanced Mass Spectrometry Facility, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, United States of America.
PLoS One. 2015 Feb 6;10(2):e0117732. doi: 10.1371/journal.pone.0117732. eCollection 2015.
Bacteroides fragilis is a widely distributed member of the human gut microbiome and an opportunistic pathogen. Cell surface molecules produced by this organism likely play important roles in colonization, communication with other microbes, and pathogenicity, but the protein composition of the outer membrane (OM) and the mechanisms used to transport polypeptides into the extracellular space are poorly characterized. Here we used LC-MS/MS to analyze the OM proteome and secretome of B. fragilis NCTC 9343 grown under laboratory conditions. Of the 229 OM proteins that we identified, 108 are predicted to be lipoproteins, and 61 are predicted to be TonB-dependent transporters. Based on their proximity to genes encoding TonB-dependent transporters, many of the lipoprotein genes likely encode proteins involved in nutrient or small molecule uptake. Interestingly, protease accessibility and biotinylation experiments indicated that an unusually large fraction of the lipoproteins are cell-surface exposed. We also identified three proteins that are members of a novel family of autotransporters, multiple potential type I protein secretion systems, and proteins that appear to be components of a type VI secretion apparatus. The secretome consisted of lipoproteins and other proteins that might be substrates of the putative type I or type VI secretion systems. Our proteomic studies show that B. fragilis differs considerably from well-studied Gram-negative bacteria such as Escherichia coli in both the spectrum of OM proteins that it produces and the range of secretion strategies that it utilizes.
脆弱拟杆菌是人类肠道微生物群中广泛分布的成员，也是一种机会致病菌。该生物体产生的细胞表面分子可能在定殖、与其他微生物的交流以及致病性方面发挥重要作用，但外膜（OM）的蛋白质组成以及将多肽转运到细胞外空间所使用的机制却鲜为人知。在这里，我们使用液相色谱-串联质谱法（LC-MS/MS）分析了在实验室条件下生长的脆弱拟杆菌NCTC 9343的外膜蛋白质组和分泌蛋白质组。在我们鉴定出的229种外膜蛋白中，预计有108种是脂蛋白，61种是依赖TonB的转运蛋白。基于它们与编码依赖TonB的转运蛋白的基因的接近程度，许多脂蛋白基因可能编码参与营养物质或小分子摄取的蛋白质。有趣的是，蛋白酶可及性和生物素化实验表明，异常大比例的脂蛋白暴露于细胞表面。我们还鉴定出三种蛋白质，它们是一个新型自转运蛋白家族成员、多个潜在的I型蛋白质分泌系统以及似乎是VI型分泌装置组成部分的蛋白质。分泌蛋白质组由脂蛋白和其他可能是推定的I型或VI型分泌系统底物的蛋白质组成。我们的蛋白质组学研究表明，脆弱拟杆菌在其产生的外膜蛋白谱和所利用的分泌策略范围方面与深入研究的革兰氏阴性菌如大肠杆菌有很大不同。
