MOLECULAR CHARACTERIZATION OF THE CLUMPING FACTOR (FIBRINOGEN RECEPTOR) OF STAPHYLOCOCCUS-AUREUS
MOLECULAR CHARACTERIZATION OF THE CLUMPING FACTOR (FIBRINOGEN RECEPTOR) OF STAPHYLOCOCCUS-AUREUS
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DOI：
10.1111/j.1365-2958.1994.tb00304.x
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发表时间：
1994-01-01
影响因子：
3.6
通讯作者：
FOSTER, TJ
中科院分区：
文献类型：
--
作者：
MCDEVITT, D;FRANCOIS, P;FOSTER, TJ
通过转座子Tn 917诱变，分离了金黄色葡萄球菌菌株纽曼的四种纤维蛋白原受体（凝集因子）缺陷突变体。突变体的Southern杂交分析鉴定了转座子-宿主DNA连接片段，其中一个片段被克隆并用于产生探针以鉴定和克隆野生型聚集因子基因座（clfA）。突变体未能在可溶性纤维蛋白原中形成团块，并且粘附在涂有纤维蛋白原的聚甲基丙烯酸甲酯（PMMA）盖玻片上。一个单拷贝的clfA基因，当引入到染色体的突变株，完全补充了这些菌株的凝集缺陷，并恢复这些突变体的能力，坚持纤维蛋白原包被的PMMA。此外，在穿梭质粒上克隆的clfA基因允许弱凝集菌株8325-4形成具有与野生型菌株纽曼相同亲合力的凝集块，并且还显著增强了8325-4菌株的粘附性。因此，可溶性纤维蛋白原中团块的形成与细菌对固相纤维蛋白原的粘附相关。clfA基因编码表观分子量为c. 130 kDa。从DNA序列推导出蛋白质的氨基酸序列;预测将表达896个残基的蛋白质（分子量92 kDa）。推定的ClfA蛋白具有表明其与细胞表面相关的特征。此外，它含有一个新的308残基区域，包括二肽重复主要是天冬氨酸和丝氨酸结束28个残基上游的LPXTG基序常见的壁相关蛋白。ClfA蛋白与S.金黄色葡萄球菌，特别是在N-和C-末端。
Four mutants of Staphylococcus aureus strain Newman that were defective in the fibrinogen receptor (clumping factor) were isolated by transposon Tn917 mutagenesis. Southern hybridization analysis of the mutants identified transposon-host DNA junction fragments, one of which was cloned and used to generate a probe to identify and clone the wild-type clumping factor locus (clfA). The mutants failed to form clumps in soluble fibrinogen and adhered poorly to polymethylmethacrylate (PMMA) coverslips coated with fibrinogen. A single copy of the clfA gene, when introduced into the chromosome of the mutant strains, fully complemented the clumping deficiency of these strains and restored the ability of these mutants to adhere to fibrinogen-coated PMMA. In addition, the cloned clfA gene on a shuttle plasmid allowed the weakly clumping strain 8325-4 to form clumps with the same avidity as the wild-type strain Newman and also significantly enhanced the adherence of 8325-4 strains. Thus the formation of clumps in soluble fibrinogen correlated with adherence of bacteria to solid-phase fibrinogen. The clfA gene encodes a fibrinogen-binding protein with an apparent molecular mass of c. 130 kDa. The amino acid sequence of the protein was deduced from the DNA sequence; it was predicted that a 896 residue protein (molecular mass 92 kDa) would be expressed. The putative ClfA protein has features that suggest that it is associated with the cell surface. Furthermore it contains a novel 308 residue region comprising dipeptide repeats predominantly of Asp and Ser ending 28 residues upstream from the LPXTG motif common to wall-associated proteins. Significant homology was found between the ClfA protein and the fibronectin-binding proteins of S. aureus, particularly in the N- and C-termini.