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首页> 外文期刊>Infection and immunity >Preparation of a sialic acid-binding protein from Streptococcus mitis KS32AR.
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Preparation of a sialic acid-binding protein from Streptococcus mitis KS32AR.

机译：从微生物链球菌KS32AR制备唾液酸结合蛋白。

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A recent report has identified a lectin on the surfaces of several strains of Streptococcus mitis and Streptococcus sanguis with specificity for an N-acetylneuraminic acid alpha 2,3-galactose-beta 1,3-N-acetylgalactosamine sequence (P.A. Murray, M.J. Levine, L.A. Tabak, and M.S. Reddy, Biochem. Biophys. Res. Commun. 106:390-396, 1982). In the present study, purification and characterization of this sialic acid-binding protein (SABP) was begun. A clinical isolate of S. mitis was grown to mid stationary phase in synthetic FMC medium and then extracted with lithium 3,5-diiodosalicylate. Lyophilized extract was subjected to gel filtration on a Sephadex G-200 column, giving four protein peaks (A to D). Peak B, shown by hemagglutination assay to contain SABP, was next subjected to affinity chromatography on a Sepharose-4B matrix coupled to fetuin glycopeptides. After an extensive washing, peak B materials bound to the affinity matrix were eluted with buffered N-acetylneuraminic acid. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis with 2-mercaptoethanol on 7.5% gels of affinity-purified materials revealed components of 96, 70, and 65 kilodaltons (kDa). Without reducing agent, only the 65-kDa band and materials which did not penetrate the gel were visualized, suggesting that the 96- and 70-kDa components were disulfide linked. The chemical cross-linking agent, disuccinimidyl suberate, was used to demonstrate specific interactions between the SABP preparation and [14C]fetuin glycopeptides. After cross-linking, sodium dodecyl sulfate-polyacrylamide gel electrophoresis and fluorography revealed the 96- and 70-kDa components, indicating that the SABP is at least bivalent. These findings support our previous suggestion that human salivary glycoproteins facilitate clearance of selected oral streptococci via specific interactions between sialic acid-containing oligosaccharides and a carbohydrate-binding protein on the bacterial cell surface.
机译：最近的一份报告已经鉴定了几种链球菌属微生物和血链球菌菌株表面的凝集素，它们对N-乙酰神经氨酸α2,3-半乳糖-β1,3-N-乙酰半乳糖胺序列具有特异性（PA Murray，MJ Levine， LA Tabak和MS Reddy，《生物化学与生物物理学》，第106卷：390-396页，1982年）。在本研究中，此唾液酸结合蛋白（SABP）的纯化和表征已开始。在合成的FMC培养基中，将临床分离的链霉菌生长至固定中期，然后用3,5-二碘水杨酸锂提取。将冻干的提取物在Sephadex G-200柱上进行凝胶过滤，得到四个蛋白质峰（A至D）。通过血凝测定法显示的峰B含有SABP，然后在与胎球蛋白糖肽偶联的Sepharose-4B基质上进行亲和层析。充分洗涤后，用缓冲的N-乙酰神经氨酸洗脱与亲和基质结合的峰B物质。用2-巯基乙醇在7.5％亲和纯化材料的凝胶上进行十二烷基硫酸钠-聚丙烯酰胺钠凝胶电泳，得到96、70和65千道尔顿（kDa）的组分。如果没有还原剂，则只能看到65 kDa的条带和未穿透凝胶的物质，这表明96 kDa和70 kDa的组分是二硫键连接的。化学交联剂辛二酸二琥珀酰亚胺酯用于证明SABP制剂与[14C] fetuin糖肽之间的特异性相互作用。交联后，十二烷基硫酸钠-聚丙烯酰胺凝胶电泳和荧光检查显示96 kDa和70 kDa组分，表明SABP至少是二价的。这些发现支持了我们先前的建议，即人类唾液糖蛋白通过细菌细胞表面上含唾液酸的寡糖和碳水化合物结合蛋白之间的特异性相互作用，促进清除选定的口腔链球菌。

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