Department of NanoBiotechnology, NanoGlycobiology, Vienna Institute of BioTechnology, Universität für Bodenkultur Wien, Muthgasse 11, A-1190 Vienna, Austria.
Department of Chemistry, Vienna Institute of BioTechnology, Universität für Bodenkultur Wien, Muthgasse 18, A-1190 Vienna, Austria.
J Biol Chem. 2011 Nov 4;286(44):38714-38724. doi: 10.1074/jbc.M111.284893. Epub 2011 Sep 12.
Cell surface glycosylation is an important element in defining the life of pathogenic bacteria. Tannerella forsythia is a Gram-negative, anaerobic periodontal pathogen inhabiting the subgingival plaque biofilms. It is completely covered by a two-dimensional crystalline surface layer (S-layer) composed of two glycoproteins. Although the S-layer has previously been shown to delay the bacterium's recognition by the innate immune system, we characterize here the S-layer protein O-glycosylation as a potential virulence factor. The T. forsythia S-layer glycan was elucidated by a combination of electrospray ionization-tandem mass spectrometry and nuclear magnetic resonance spectroscopy as an oligosaccharide with the structure 4-Me-β-ManpNAcCONH(2)-(1→3)-[Pse5Am7Gc-(2→4)-]-β-ManpNAcA-(1→4)-[4-Me-α-Galp-(1→2)-]-α-Fucp-(1→4)-[-α-Xylp-(1→3)-]-β-GlcpA-(1→3)-[-β-Digp-(1→2)-]-α-Galp, which is O-glycosidically linked to distinct serine and threonine residues within the three-amino acid motif (D)(S/T)(A/I/L/M/T/V) on either S-layer protein. This S-layer glycan obviously impacts the life style of T. forsythia because increased biofilm formation of an UDP-N-acetylmannosaminuronic acid dehydrogenase mutant can be correlated with the presence of truncated S-layer glycans. We found that several other proteins of T. forsythia are modified with that specific oligosaccharide. Proteomics identified two of them as being among previously classified antigenic outer membrane proteins that are up-regulated under biofilm conditions, in addition to two predicted antigenic lipoproteins. Theoretical analysis of the S-layer O-glycosylation of T. forsythia indicates the involvement of a 6.8-kb gene locus that is conserved among different bacteria from the Bacteroidetes phylum. Together, these findings reveal the presence of a protein O-glycosylation system in T. forsythia that is essential for creating a rich glycoproteome pinpointing a possible relevance for the virulence of this bacterium.
细胞表面糖基化是定义致病菌生命的重要因素。坦纳拉福赛思氏菌是一种革兰氏阴性、厌氧性牙周致病菌，栖息在下牙龈菌斑生物膜中。它完全被由两种糖蛋白组成的二维结晶表面层 (S-层) 覆盖。尽管先前已经表明 S-层可以延缓细菌被先天免疫系统识别，但我们在这里将 S-层蛋白 O-糖基化特征化为潜在的毒力因子。T. forsythia S-层聚糖通过电喷雾串联质谱和核磁共振波谱学的组合来阐明，其结构为寡糖，结构为 4-Me-β-ManpNAcCONH(2)-(1→3)-[Pse5Am7Gc-(2→4)-]-β-ManpNAcA-(1→4)-[4-Me-α-Galp-(1→2)-]-α-Fucp-(1→4)-[-α-Xylp-(1→3)-]-β-GlcpA-(1→3)-[-β-Digp-(1→2)-]-α-Galp，通过 O-糖苷键与 S-层蛋白上三个氨基酸基序 (D)(S/T)(A/I/L/M/T/V) 中独特的丝氨酸和苏氨酸残基相连。这种 S-层聚糖显然会影响 T. forsythia 的生活方式，因为 UDP-N-乙酰氨基葡萄糖醛酸脱氢酶突变体的生物膜形成增加可以与截短的 S-层糖链的存在相关。我们发现 T. forsythia 的几种其他蛋白质也被该特定寡糖修饰。蛋白质组学鉴定出其中两种属于先前分类的抗原性外膜蛋白，这些蛋白在生物膜条件下上调，此外还有两种预测的抗原性脂蛋白。对 T. forsythia 的 S-层 O-糖基化的理论分析表明，存在一个 6.8kb 的基因座，该基因座在杆菌门的不同细菌中保守。总之，这些发现揭示了 T. forsythia 中存在蛋白质 O-糖基化系统，该系统对于创建丰富的糖蛋白组至关重要，这可能表明该细菌的毒力具有相关性。
