DR ADHESINS IN E COLI RENAL VS BLADDER TROPISM

表达Dr粘连素的大肠杆菌引起肾盂肾炎，相关的AFA-III引起膀胱炎。本研究旨在确定DraE的受体结合位点，并为肾向性和膀胱向性的机制提供见解。目的1。鉴定和表征与DAF相互作用的Dr菌膜结构域。我们提出DraE的n端表面暴露残基形成一个结合DAF的组装表位。亲水性结构域Val30至Pro42将通过丙氨酸扫描诱变发生突变，并分析突变体对DAF+ CHO细胞的结合和侵袭。目标2。目的：探讨Dr+大肠杆菌嗜肾性与嗜膀胱性的致病机制。DraE和AWE在三个氨基酸（D54N, T90M, I113T）上有所不同。博士+ E。大肠杆菌感染肾脏，而fa - iii +大肠杆菌感染膀胱。将DraE的氨基酸替换为Afa3E的氨基酸，并用大肠杆菌HI 11128的等基因突变体分析小鼠的肾性。目标3。目的：研究DraE和Afa3E黏附素对细胞的附着和侵袭。DraE比Afa3E更具侵入性。我们将用涂有纯化的DraE突变体的聚苯乙烯球，免疫金染色和电子显微镜研究入侵。解决结合区域将使肽抑制剂设计针对共同受体，防止细菌分散和细胞入侵。

Escherichia coli expressing Dr adhesin cause pyelonephritis and related AFA-III cause cystitis. This study seeks to identify the receptor-binding site of DraE and provide insights into the mechanism of renal vs bladder tropism. Aim 1. To identify and characterize domains of Dr fimbriae that interact with DAF. We propose that N-terminal surface exposed residues of DraE form an assembled epitope that bind DAF. The hydrophilic domain Val30 to Pro42 will be mutated by alanine-scanning mutagenesis and mutants analyzed for binding and invasion of DAF+ CHO cells. Aim 2. To investigate the pathogenic mechanism that determines kidney vs bladder tropism of Dr+ E. coli. DraE and AWE differ in three amino acids (D54N, T90M, I113T). Dr+E. coli infect the kidneys while AFA-III+ E. coli infect the bladder. Amino acids of DraE will be replaced with those of Afa3E and renal tropism analysed using isogenic mutants of E. coli HI 11128 in mice. Aim 3. To study attachment and cell invasion by DraE and Afa3E adhesins. DraE is more invasive than Afa3E. We will study invasion using polystyrene beads coated with purified DraE mutants, immunogold staining and electron microscopy. Resolving the binding domain will enable design of peptide inhibitors against the common receptor preventing bacterial dispersion and cell invasion.