IDENTIFICATION OF A STAPHYLOCOCCUS-AUREUS EXTRACELLULAR MATRIX-BINDING PROTEIN WITH BROAD SPECIFICITY
IDENTIFICATION OF A STAPHYLOCOCCUS-AUREUS EXTRACELLULAR MATRIX-BINDING PROTEIN WITH BROAD SPECIFICITY
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DOI：
10.1128/iai.61.6.2479-2485.1993
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发表时间：
1993-06-01
影响因子：
3.1
通讯作者：
HOOK, M
中科院分区：
文献类型：
--
作者：
MCGAVIN, MH;KRAJEWSKAPIETRASIK, D;HOOK, M
我们试图鉴定金黄色葡萄球菌细胞上骨唾液蛋白（BSP）的受体，从而纯化了一种能够结合几种细胞外基质糖蛋白的葡萄球菌表面蛋白。将不同葡萄球菌菌株的蛋白溶解于十二烷基硫酸钠中，聚丙烯酰胺凝胶电泳分离，印迹于Immobilon P膜上，并用I-125-BSP探针检测。几种细菌蛋白结合放射性标记的配体，不同的菌株表达不同的bsp结合蛋白。在大多数菌株中存在表观M(r)s为72,000或60,000的主要bsp结合蛋白，并对这些蛋白进行了进一步研究。细菌在Luria培养液中比在tryptic大豆培养液中优先表达72- kda和60-kDa蛋白，其丰度可能与I-125-BSP结合增加有关。72-kDa和60-kDa蛋白均通过1 M LiCl萃取细胞溶解，并通过阳离子交换层析纯化。对纯化的72kda蛋白的氨基酸组成分析表明，该蛋白含有较高的赖氨酸（11.9%）和疏水氨基酸（28.0%）。在Western配体印迹（免疫印迹）实验中，72-kDa蛋白不仅结合BSP，还结合放射标记的纤维连接蛋白、纤维蛋白原、玻璃体连接蛋白、血栓反应蛋白，在某种程度上还结合胶原蛋白。将纯化的60 kda蛋白添加到金黄色葡萄球菌细胞中并没有抑制不同配体的结合，但在某些情况下导致i -125配体的结合增强。纯化的60kda蛋白在61马克杯/毫升的浓度下能使绵羊红细胞发生血凝。高浓度的焦糖、甘露糖或糖二糖可抑制凝集反应。这些数据表明，纯化后的蛋白可能作为细菌受体，对基质糖蛋白具有广泛的特异性，并且可能作为碳水化合物结合蛋白。
A staphylococcal surface protein capable of binding several extracellular matrix glycoproteins was purified as a result of our attempts to identify a receptor(s) for bone sialoprotein (BSP) on Staphylococcus aureus cells. Proteins from different staphylococcal strains were solubilized in sodium lauryl sulfate, separated by polyacrylamide gel electrophoresis, blotted onto Immobilon P membranes, and probed with I-125-BSP. Several bacterial proteins bound the radiolabeled ligand, and various strains expressed different repertoirs of BSP-binding proteins. Major BSP-binding proteins with apparent M(r)s of 72,000 or 60,000 were present on most strains, and these proteins were further studied. The 72- and 60-kDa proteins were preferentially expressed when bacteria were cultured in Luria broth compared with when they were cultured on tryptic soy broth, and the abundance of the proteins could be correlated to an increased I-125-BSP binding. Both the 72-kDa and the 60-kDa proteins were solubilized by extraction of cells with 1 M LiCl and were purified by cation-exchange chromatography. Amino acid composition analysis of the purified 72-kDa protein indicated a high content of lysine (11.9%) and hydrophobic amino acids (28.0% combined). In Western ligand blotting (immunoblotting) experiments, the 72-kDa protein bound not only BSP but also radiolabeled fibronectin, fibrinogen, vitronectin, thrombospondin, and, to some extent, collagen. Addition of the purified 60-kDa protein to S. aureus cells did not inhibit binding of the different ligands but in some cases resulted in an augmentation of the binding of I-125-ligand. Purified 60-kDa protein could hemagglutinate sheep erythrocytes at a concentration of 61 mug/ml. The agglutination reaction was inhibited by high concentrations of fucose, mannose, or melibiose. These data suggest that the purified proteins may serve as bacterial receptors with broad specificity for matrix glycoproteins and thal the proteins may act as carbohydrate-binding proteins.