PHOSPHATASE AS A VIRULENCE FACTOR IN Q FEVER

磷酸酶作为 Q 热的毒力因子

• 批准号: 2067386
• 负责人: OSWALD G BACA
• 金额: $ 19.29万
• 依托单位: UNIVERSITY OF NEW MEXICO
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-04-01 至 1996-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2067386
• 关键词: Q fever; SDS polyacrylamide gel electrophoresis; acid phosphatase; active sites; density gradient ultracentrifugation; enzyme activity; enzyme linked immunosorbent assay; enzyme substrate; gene complementation; high performance liquid chromatography; host organism interaction; human subject; leukocyte oxidative burst; molecular cloning; molecular pathology; neutrophil; phagocytes; phosphoprotein phosphatase; protein sequence; scintillation counter; transmission electron microscopy; virulence; western blottings

Coxiella burnetii, the rickettsial agent of Q fever in humans, is an obligate intracellular bacterium. Occasionally, acute disease is followed by life-threatening chronic endocarditis for which there is no consistently effective treatment. The long-term goal of this research is to discover the biochemical/molecular mechanisms that account for survival of C burnetii in the phagolysosomes of phagocytic cells and other host cells. Preliminary studies have revealed that the organism possesses significant acid phosphatase activity and that it may be responsible for inhibiting the oxidative burst in human neutrophils; this indicates that the enzyme may be a major virulence determinant. Phosphatases have been shown to occur at or near the surface of Leishmania promastigotes and Legionella micdadei, a bacterium that is phylogenetically related to C burnetii. The phosphatases inhibit the oxidative burst and concomitant production of toxic superoxide anion by phagocytosing human neutrophils; this probably accounts, in large part, for the survival of these parasites within phagocytes. The specific aims of the proposed research are to: (i) characterize the biochemical and biophysical properties of the recently discovered periplasmic C burnetii acid phosphatase; (ii) clone and characterize the acid phosphatase gene using available gene banks; (iii) examine the effect of the enzyme on human neutrophils. Preliminary evidence strongly indicates that partially purified C burnetii acid phosphatase inhibits the oxidative burst of human neutrophils; this will be verified with purified phosphatase. The site and mechanism of action of the enzyme will be investigated. Whether or not the phosphatase is released from the parasite during the infection process will be determined. These aims will provide basic information about an enzyme which may play a key role in blocking the host cell's capacity to mount a response against the invading and resident parasites. Attainment of these goals will result in a better understanding of the parasite's capacity to circumvent the host's defenses during invasion of and subsequent survival and proliferation within phagocytes and other host cells. Such knowledge is essential for devising strategies for the development of efficacious drugs and vaccines for treating/preventing Q fever, including life- threatening endocarditis.

伯氏柯克斯体是人类Q热的立克次体病原体，是一种 专属胞内细菌。偶尔，急性疾病是 其次是危及生命的慢性心内膜炎，目前还没有 一贯有效的治疗方法。这项研究的长期目标是 就是发现生物化学/分子机制 伯氏梭菌在吞噬细胞吞噬溶酶体中的存活和 其他宿主细胞。初步研究表明，这种有机体 具有显著的酸性磷酸酶活性，它可能是 负责抑制人类中性粒细胞的氧化爆发；这 表明该酶可能是一个主要的毒力决定因素。 磷酸酶已被证明存在于表面或接近表面 利什曼原虫和米氏军团菌，一种 在系统发育上与伯内提毛虫有亲缘关系。磷酸酶抑制了 氧化性猝发及伴随产生的有毒超氧阴离子 吞噬人类中性粒细胞；这可能在很大程度上解释了， 为了这些寄生虫在吞噬细胞中的生存。具体目标 拟议研究的目的是：(I)描述生物化学和 新近发现的周质伯氏杆菌的生物物理性质 酸性磷酸酶；(Ii)酸性磷酸酶基因的克隆和鉴定 利用现有的基因库；(Iii)检查该酶对 人类中性粒细胞。初步证据有力地表明 部分纯化的伯氏杆菌酸性磷酸酶抑制氧化 人类中性粒细胞爆裂；这将通过纯化得到验证 磷酸酶。该酶的作用部位和作用机制将是 调查过了。无论磷酸酶是否从细胞中释放 将确定感染过程中的寄生虫。这些目标 将提供有关一种可能起关键作用的酶的基本信息 在阻止宿主细胞对 入侵和寄生的寄生虫。这些目标的实现将会带来 为了更好地了解寄生虫绕过 寄主在入侵和随后存活期间的防御 吞噬细胞和其他宿主细胞内的增殖。这样的知识是 对于制定有效的发展战略至关重要 治疗/预防Q热的药物和疫苗，包括生命 有威胁的心内膜炎。