SECRETION OF PERTUSSIS TOXIN FROM BORDETELLA PERTUSSIS

百日咳博德特氏菌分泌百日咳毒素

• 批准号: 2568889
• 负责人: D L BURNS
• 金额: --
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/2568889
• 关键词: Bordetella pertussis; bacterial genetics; bacterial proteins; gene expression; gene interaction; immunoprecipitation; mutant; pertussis toxin; protein purification; secretion; secretory protein; transport proteins

Previously, we demonstrated the existence of a set of eight genes that are essential for the secretion of pertussis toxin (PT) from B. pertussis. These genes, termed ptl genes, encode transport proteins that share homology with a family of proteins involved in the transport of both DNA and proteins across bacterial membranes. We now have evidence for a ninth secretion gene. This small gene, ptl1 is predicted to encode a protein of 6.8 kDa and is located between ptlD and ptlE. PtlI shares homology with VirB7 which is required for transport of T-DNA across the membranes of the plant pathogen Agrobacterium tumefaciens. Deletion of the ptlI region from B. pertussis has no discernable effect of PtlE but resulted in loss of detectable PtlF, suggesting the PtlI may interact with and stabilize PtlF. Using immunoprecipitation techniques, we demonstrated a direct interaction between PtlI and PtlF. These results are the first evidence of interactions between Ptl proteins. In addition to the work on elucidation of interactions between Ptl proteins, we are also investigating the mechanism by which the Ptl proteins promote transport of PT. One Ptl protein in particular, PtlC, has the interesting characteristic that it contains a nucleotide binding motif. We are currently purifying this protein and determining whether this protein has either hydrolase or kinase activity that might play a critical role in the transport process. This work increases our knowledge of the secretion of PT and may aid in the construction of strains of B. pertussis which efficiently produce and secrete PT. Such strains would be useful for vaccine production. In addition, knowledge of the mechanism of PT secretion might aid in the development of live-attenuated vaccines that secrete inactivated forms of PT, thus inducing a protective immune response.

在此之前，我们演示了一组由八个基因组成的 是B。百日咳毒素(PT)分泌所必需的。 百日咳。这些基因被称为PTL基因，编码的运输蛋白 与参与运输的一组蛋白质具有同源性 DNA和蛋白质都能穿过细菌的膜。我们现在有证据 为了第九个分泌基因。这种名为ptl1的小基因被预测为编码 蛋白质大小为6.8 kDa，位于pTLD和ptlE之间。PTLI股份 与病毒B7同源，病毒B7是T-DNA跨 植物病原菌根癌农杆菌的膜。删除 百日咳杆菌的ptlI区没有ptlE的明显作用，但 导致可检测到的PtlF的丢失，表明PtlI可能相互作用 并稳定PtlF。使用免疫沉淀技术，我们 证实了PtlI和PtlF之间的直接相互作用。这些结果 是PTL蛋白之间相互作用的第一个证据。此外 对于阐明PTL蛋白之间相互作用的工作，我们是 也研究了PTL蛋白促进的机制 PT的运输。一种特别的Ptl蛋白，PtlC，具有 有趣的特征是它包含一个核苷酸结合基序。 我们目前正在提纯这种蛋白质，并确定这是否 蛋白质具有水解酶或激酶活性，可能起到 在运输过程中发挥关键作用。这项工作增加了我们的 对甲状旁腺素分泌的了解，并可能有助于构建 高效产生和分泌PT的百日咳杆菌菌株。是这样的 菌株将对疫苗生产有用。此外，知识 凝血酶原激活剂分泌机制的研究可能有助于 分泌灭活的PT形式的减毒活疫苗，因此 诱导保护性免疫反应。