Investigation for regulatory mechanisms of GcrR protein for S.mutans gbp Cexpression

作者研究了gcrR基因作为gbpc基因表达双组分调控系统的反应调节因子的调控机制。反应调节蛋白的信号转导是通过其相应的传感器-递质蛋白的磷酸化转移来完成的。通过定点突变，将GcrR蛋白的第53个氨基酸天冬氨酸作为信号受体位点改变为花生氨酸，并将该突变引入到变形链球菌染色体中。尽管该突变体中的这种改变的GcrR蛋白不应该被磷酸化，但该突变体表现出与野生型菌株相同的葡聚糖依赖的聚集表型。因此，作者得出结论，GcrR蛋白不是作为反应调节蛋白，而是作为抑制蛋白。Gbpc蛋白是一种与肽聚糖结合的壁锚式蛋白。Western印迹分析证实gbpc在细胞壁有表达，表明变形链球菌细胞具有…功能。利用Biacore系统，更多地结合到gbpc蛋白介导的固定化葡聚糖上。蛋白质的细胞壁锚定是由索酸酶介导的。变形链球菌的山梨糖酶阴性突变株没有表现出预期的葡聚糖依赖的聚集表型。在菌株GS-5中首次发现gbpc基因的无义突变，这导致在变形链球菌中检测到gbpc基因的单核苷酸多态。Gbpc基因的保守区和多态区均可用于gbpc蛋白功能域的估计和变形链球菌的鉴定。根据这一保守的gbpc蛋白信息设计了一对引物，成功地用PCR方法鉴定了猕猴链球菌gbpc基因的同源物。作者发现了另一种被命名为冷凝集的聚集表型。采用Himarl转座子体外转座的随机诱变方法，从部分变形链球菌中鉴定了胶原结合粘附素基因。这是第一份在人类口腔本土菌群中证明了来自绿色链球菌的胶原结合粘附素的报告。较少

The auther has investigated the regulatory mechanisms of the gcrR gene as a response regulator of two-component regulatory systems for gbpC gene expression. Signal transduction of response regulators is performed by their phosphoryl transfer from their counterparts, sensor-transmitter proteins. The 53rd amino acid aspartate of GcrR protein that is expected as the signal receptor site was changed to aranine by site-directed mutagenesis, and this mutation was introduced into S.mutans chromosome. Albeit this altered GcrR protein in this mutant should not be phosphorylated, this mutant exhibited the same dextran-dependent aggregation phenotype as the wild type strain. Therefore, the authers concluded that GcrR protein does not act as a response regulator but act as a represser. GbpC protein is a member of wall-anchored proteins tethered to peptidoglycan layer. The authers confirmed GbpC expression at cell wall by the Western blot analysis, and the S.mutans cells were indicated to be able t … More o bind to immobilized glucan mediated by the GbpC protein using the BiaCore system. Cell wall anchoring of the protein is mediated by the sortase enzyme. Sortase negative mutant of S.mutans did not exhibit dextran-dependent aggregation phenotype as expected. A nonsense mutation of the gbpC gene was initially found in strain GS-5 and this lead to detection of single nucleotide polymorphisms in the gbpC gene among S.mutans strains. Both conserved and polymorphic regions of the gbpC gene will be useful for an estimation of functional domains of GbpC protein and identification of S.mutans strains. A gbpC gene homologue from Streptococcus macacae was successfully identified by PCR method with primers designed from this conserved GbpC protein information. The authers found another aggregation phenotype designated as cold agglutination. The collagen-binding adhesin gene was identified from some strains of S.mutans by random mutagenesis method using in vitro transposition of Himarl transposon. This is the first report that demonstrates a collagen-binding adhesin from viridans streptococci in human oral indigenous flora. Less