Murray P A, Levine M J, Reddy M S, Tabak L A, Bergey E J
Infect Immun. 1986 Aug;53(2):359-65. doi: 10.1128/iai.53.2.359-365.1986.
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.
最近的一份报告指出，在几种缓症链球菌和血链球菌菌株的表面存在一种凝集素，它对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（P.A. 默里、M.J. 莱文、L.A. 塔巴克和M.S. 雷迪，《生物化学与生物物理研究通讯》106:390 - 396，1982年）。在本研究中，开始了对这种唾液酸结合蛋白（SABP）的纯化和特性鉴定。将一株缓症链球菌临床分离株在合成FMC培养基中培养至对数中期，然后用3,5 - 二碘水杨酸锂进行提取。冻干提取物在Sephadex G - 200柱上进行凝胶过滤，得到四个蛋白峰（A至D）。通过血凝试验显示峰B含有SABP，接下来将其在与胎球蛋白糖肽偶联的Sepharose - 4B基质上进行亲和层析。经过大量洗涤后，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。在7.5%凝胶上用2 - 巯基乙醇进行十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳，对亲和纯化的物质进行分析，结果显示有96、70和65千道尔顿（kDa）的组分。在没有还原剂的情况下，仅可见65 - kDa条带和未穿透凝胶的物质，这表明96 - kDa和70 - kDa组分通过二硫键相连。化学交联剂辛二酸二琥珀酰亚胺酯用于证明SABP制剂与[14C]胎球蛋白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自显影显示出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之前的推测，即人类唾液糖蛋白通过含唾液酸的寡糖与细菌细胞表面的碳水化合物结合蛋白之间的特异性相互作用，促进了特定口腔链球菌的清除。
