Structure and Function of Type IV Pili in Clostridium difficile

艰难梭菌IV型菌毛的结构和功能

• 批准号: 8811000
• 负责人: Kurt Henry Piepenbrink
• 金额: $ 5.6万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-02-01 至 2016-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8811000
• 关键词: Adherence; Bacillus (bacterium); Bacteria; Bacteriology; Basic Science; Binding; Biogenesis; Biological Assay; Calorimetry; Cell Adhesion; Cells; Clinical; Clostridium; Clostridium difficile; Clostridium perfringens; Colitis; Collaborations; Colon; Crystallography; Cysteine; Data; Deuterium; Diarrhea; Electrons; Fiber; Fimbriae Proteins; Gene Cluster; Genes; Genome; Goals; Gram-Negative Bacteria; Gram-Positive Bacteria; Hair; Health; Horizontal Gene Transfer; Human; Hydrogen; Immune; Immune response; Immune system; Incidence; Infection; Investigation; Knowledge; Label; Learning; Maps; Mass Spectrum Analysis; Measures; Mediating; Microbial Biofilms; Modeling; Molecular; Molecular Conformation; NMR Spectroscopy; Organism; Peptides; Physiological; Pilum; Play; Prevention approach; Process; Protein Subunits; Proteins; Pseudomembranous Colitis; Publishing; Reagent; Relaxation; Reproduction spores; Resolution; Role; Ruminococcus; Sampling; Severities; Signal Transduction; Source; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Structure; Surface; Surface Plasmon Resonance; Technetium Tc 99m ciprofloxacin; Techniques; Tissues; Titrations; Toxic Megacolon; Toxic effect; Vaccines; Virulence; X-Ray Crystallography; antimicrobial; appendage; cell motility; crosslink; disulfide bond; gastrointestinal; insight; member; monomer; mortality; novel; novel strategies; novel vaccines; prevent; receptor

DESCRIPTION (provided by applicant): Clostridium difficile, a spore-forming anaerobic Gram-positive bacillus, is the cause of a spectrum of gastrointestinal illness ranging from mild diarrhe though pseudomembranous colitis, and toxic megacolon. Alarmingly, the incidence, severity and mortality of C. difficile colitis have all increased significantly in the past twenty years. Th mechanism of C. difficile toxicity is well-characterized but no vaccine against C. difficile infecton exists and our knowledge about the interactions of C. difficile with its host is limited. One possible target for the host immune system are the Type IV pili of C. difficile. Type IV Pili (T4P) are hair-like surface appendages produced by many species of pathogenic Gram negative bacteria which play a role in diverse processes such as cellular adhesion, colonization, twitching motility, biofilm formation, horizontal gene transfer and in numerous instances are essential for virulence. T4Ps are composed exclusively or primarily of many copies of pilin protein, tightly packed in a helix so that the highly hydrophobic amino-terminus of the pilin is buried in the pilus core. This project aims to characterize the structure and functional significance of the proteins that make up the Type IV pili of C. difficile. T4Ps are produced by many species of pathogenic Gram-negative bacteria and are known to play a role in both cellular adhesion and colonization. T4Ps are composed exclusively or primarily of many copies of a pilin protein, tightly packed in a helix. Although better characterized in Gram-negative bacteria, T4P genes have been discovered in the genomes of all members of the Clostridium genus, including C. difficile. In collaboration with Dr. Michael Donnenberg, we have established structural investigations into six putative pilin genes identified in the C. difficile genome and have solved the structure of one PILA4, by x-ray crystallography. Rather than consisting of one β-sheet surrounded by α-helices, as is the case with all other known pilins, PILA4 contains a second β-sheet at a ~70 angle to the helical pilin core. This second sheet and the surrounding helices give it a much more extended conformation from the core and this radically different exposed surface may play a role in cellular adhesion and possibly signaling of innate immune cells. We have used this structure in combination with data from other sources to model the structure of a pilus composed of PilA4. We have also grown isotopically-labeled samples of another pilin, PilA2, and have collected data using NMR spectroscopy to being assigning the resonances to determine the structure using distance and relaxation rates from NMR. Functional studies of PILA4 are currently underway while structural investigations into other pilin proteins continue both by x-ray crystallography and NMR.

描述（由申请人提供）：艰难梭菌，一种形成孢子的厌氧革兰氏阳性芽孢杆菌，是一系列胃肠道疾病的病因，从轻度腹泻到假膜性结肠炎，以及中毒性巨结肠。令人震惊的是，艰难梭菌结肠炎的发病率、严重程度和死亡率在过去二十年中都有显著增加。艰难梭菌的毒性机制已经很好地描述了，但目前还没有针对艰难梭菌感染的疫苗，我们对艰难梭菌与宿主相互作用的了解也很有限。宿主免疫系统的一个可能目标是艰难梭菌的IV型菌毛。IV型菌毛（T4P）是由多种致病性革兰氏阴性菌产生的毛发状表面附属物，在多种过程中发挥作用，如细胞粘附、定植、收缩运动、生物膜形成、水平基因转移，在许多情况下对毒力至关重要。T4Ps完全或主要由绒毛蛋白的许多拷贝组成，它们紧密地排列成螺旋状，因此绒毛蛋白高度疏水的氨基末端被埋在菌毛中