FIMBRIAL ADHERENCE TO LUNG EPITHELIA BY KLEBSIELLA

克雷伯菌对肺上皮细胞的菌毛粘附

• 批准号: 2210175
• 负责人: DOUGLAS B HORNICK
• 金额: $ 7.63万
• 依托单位: UNIVERSITY OF IOWA
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-04-01 至 1997-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2210175
• 关键词: Klebsiella; adhesin; alleles; antibacterial antibody; bronchoscopy; cell adhesion; cell transformation; collagen; epitope mapping; gene deletion mutation; gene mutation; human subject; human tissue; laboratory rabbit; lung; mutant; nucleic acid sequence; pilus; protein sequence; respiratory epithelium; site directed mutagenesis; trachea

Research: Attachment the respiratory epithelium is believed to be an early step in the development of pneumonia. Klebsiella produce filamentous organelles called fimbriae which carry adhesive protein subunits called adhesins. This proposal will further characterize the Klebsiella type 3 fimbrial adhesin responsible for adherence to respiratory epithelium. The initial experiments will identify which DNA sequence(s) of the type 3 fimbrial adhesin gene, mrkD, are critical for proper function of the adhesin. The two necessary recombinant plasmids producing transformed bacteria that express nonadhesive type 3 fimbriae (pFK28) and type 3 fimbrial adhesin protein (pFK58) were previously constructed. Hence, in this study, site-specific and random mutations in pFK58 can be produced and the appropriate mutations identified by DNA sequencing. The effect of the mutations on adherence by cotransformed bacteria will be quantified by three established adherence assays to type V collagen, human tracheal cells, and lung tissue sections. In subsequent experiments, type 3 fimbrial adhesin antibodies will be developed to assist in experiments to localize the adhesin on Klebsiella type 3 fimbrial filament as well as on mutant strains, developed in earlier experiments, that may demonstrate increased or decreased adherence. Additional experiments will utilize synthetic nonameric peptides, derived from the region(s) of the mrkD gene shown to be important, to determine the primary amino acid sequence required for adherence by the type 3 adhesin and to determine if these regions encompass epitopes recognized by adhesin-specific antisera. In the last group of experiments the applicant will use a novel method of allelic exchange to generate specific deletions in the Klebsiella mrk gene cluster within the native chromosomal DNA. Subsequently the impact on adherence by these specifically mutated Klebsiella will be assessed in the three adherence assays.

研究：附着于呼吸上皮被认为是一种