酒酒球菌小分子热休克蛋白sHsp20生物学功能及其作用机理研究

In this study, the biological functions and mechanisms of sHsp20 were firstly investigated. The coding sequence of sHsp20 was amplified by PCR with genome DNA as template. E. coli harboring the expression vector was screened. The protein was purified by Nickle column and anion exchange chromatography. The cell viability of E.coli with and without sHsp20 expression at different stress conditions were compared. The functions and characteristics of sHsp20 were investigated in vitro and in vivo. The structural model of sHsp20 dimer was predicted by homology modeling. On the basis of the model, the oligomeric dissociation correlated with chaperone-like activities of sHsp20 were revealed by chemical cross-linking. Key amino acid residues were mutated and cloned. The corresponding mutants were expressed and purified, and their structure and function were studied, and were compared to that of sHsp20. A better understanding of the relationship between structure and function of sHsp20 will be of theoretical and practical importance.

本项目以酒酒球菌小分子热休克蛋白sHsp20为研究对象。通过现代分子生物学技术克隆酒酒球菌小分子热休克蛋白sHsp20基因，构建大肠杆菌表达菌株，纯化得到重组蛋白sHsp20。从作用于底物蛋白和细胞膜两方面研究sHsp20在表达体内外的生物学功能和本身特性。利用生物信息学方法，通过多序列比对寻找合适的模板，同源建模得到sHsp20的三级结构模型。以此为基础，利用化学交联等方法，研究sHsp20聚体形式与活性的关系。对关键结构位点进行突变，得到突变蛋白，利用现代分析仪器研究野生型和突变蛋白的结构与功能的差异，进一步探讨sHsp20作用机理。该项目的实施，进一步丰富了sHsp的种类和功能领域的研究，为揭示sHsp结构与功能关系以及sHsp作用机理的研究提供更全面的理解，为sHsp的实际应用提供基础数据。

本项目以酒酒球菌小分子热休克蛋白sHsp20为研究对象。通过现代分子生物学技术克隆酒酒球菌小分子热休克蛋白sHsp20基因，构建大肠杆菌表达菌株，纯化得到重组蛋白sHsp20。从作用于底物蛋白和细胞膜两方面研究sHsp20在表达体内外的生物学功能和本身特性。利用生物信息学方法，通过多序列比对寻找合适的模板，同源建模得到sHsp20的三级结构模型。以此为基础，利用化学交联等方法，研究sHsp20聚体形式与活性的关系。对关键结构位点进行突变，得到突变蛋白，利用现代分析仪器研究野生型和突变蛋白的结构与功能的差异，进一步探讨sHsp20作用机理。该项目的实施，进一步丰富了sHsp的种类和功能领域的研究，为揭示sHsp结构与功能关系以及sHsp作用机理的研究提供更全面的理解，为sHsp的实际应用提供基础数据。