Amylase Binding Streptococci Dental Plaque and Caries

描述：牙菌斑是导致龋齿和牙周病的口腔生物膜。许多唾液蛋白已被证明与菌斑中的细菌相互作用，这种相互作用可能在菌斑形成中起关键作用。其中一种相互作用是淀粉酶（唾液中最丰富的酶）与淀粉酶结合链球菌（ABS）之间的相互作用，而淀粉酶结合链球菌（ABS）在菌斑中数量众多。基于我们在之前资助期间的最新发现，这个经过修改的竞争性更新申请旨在继续我们对淀粉酶与ABS结合的生化、生理学、龋齿学和生态学后果的研究。我们发现，尽管淀粉酶结合缺陷突变体在淀粉酶包被表面的粘附性较差，并且在体外表现出有缺陷的生物膜形成，但它们比野生型菌株更能在大鼠牙齿上定植。在喂食蔗糖/淀粉食物的大鼠中胜过亲本品系。这些令人惊讶的发现使人们认识到，AbpA抑制了依赖蔗糖的定植决定因素，例如，但可能不限于，S. gordonii的GtfG。因此，我们下一个资助期的目标是：1)研究参与淀粉酶结合的基因和蛋白质（abpA和abpB）与参与葡聚糖合成的基因（rgg和gtfG）之间的潜在相互作用。我们将评估突变菌株中abpA、abpB和gtfG的转录，以确定Abp是否在转录水平上调节gtfG的表达（反之亦然）。2)比较野生型和突变型菌株在标准体外粘附和生物膜模型中的差异。3)利用蛋白质组学方法评估Gtf和淀粉酶结合蛋白之间的物理相互作用，如Western blot-配体结合试验、Maldi-Tof（基质辅助、激光解吸电离/飞行时间）质谱法或噬菌体展示。4)确定gtf缺乏或精通的哥氏弧菌的淀粉酶结合突变是否会调节哥氏弧菌在大鼠中的口腔定植和致龋性。5)确定戈多氏葡萄球菌-淀粉酶相互作用是否调节变形葡萄球菌的定植竞争和致龋性，以及表达AbpA的变形葡萄球菌菌株是否表现出改变的定植和/或致龋能力。结合体内和体外的研究对于了解牙菌斑形成的机制至关重要。

DESCRIPTION: Dental plaque is the oral biofilm responsible for the etiology of Dental caries and periodontal disease. A number of salivary proteins have been shown to interact with bacteria in plaque, and such interactions likely play critical roles in plaque formation. One such interaction is that between amylase, the most abundant enzyme in saliva, and the amylase-binding streptococci (ABS), which are numerous in plaque. Based on our recent findings generated during the previous funding period, this once-amended competitive renewal application seeks to continue our studies of the biochemical, physiological, cariological and ecological consequences of amylase-binding to ABS. We found that while amylase binding-deficient mutants adhere less well to amylase-coated surfaces and demonstrate defective biofilm formation in vitro, they colonize rat teeth better than wild type strains, out-compete their parental strains in rats fed sucrose/starch diet. These surprising findings led to the realization that AbpA inhibits sucrose-dependent colonization determinants such as, but perhaps not limited to, GtfG of S. gordonii. Thus, our Aims for the next funding period are to: 1) investigate potential interactions of genes and proteins involved in amylase binding (abpA and abpB) and genes involved in glucan synthesis (rgg and gtfG). We will evaluate the transcription of abpA, abpB and gtfG in mutant strains to determine if Abp modulates gtfG expression (or vice versa) at the transcriptional level. 2) compare wildtype and mutant strains in standard in vitro adhesion and biofilm models. 3) assess the physical interaction between Gtf and amylase-binding proteins using proteomic approaches such as Western blot-ligand binding assays, Maldi-Tof (Matrix-assisted, Laser-Desorption-Ionization/Time of Flight) mass spectrometry, or phage display. 4) determine if amylase-binding mutations in Gtf-deficient or -proficient S. gordonii modulate S. gordonii oral colonization and cariogenicity in rats. 5) determine if S. gordonii-amylase interactions modulate S. mutans colonization competition and cariogenicity and if strains of S. mutans made to express AbpA show altered colonization and/or cariogenic abilities. Integration of in vitro with in vivo studies is crucial for mechanistic understanding of Dental plaque formation.