用蛋白电泳和高效凝胶排阻色谱法分析还原脲变性蛋白溶菌酶稀释复性过程的集聚体

本文利用蛋白电泳和高效凝胶排阻层析法分析了还原脲变性蛋白溶菌酶稀释复性过程中的集聚体。当用复性液稀释复性还原脲变性蛋白溶菌酶时,会迅速产生可观量的沉淀。沉淀和上清液的不连续十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)和高效凝胶排阻层析分析结果表明,还原脲变性蛋白溶菌酶在稀释复性过程中除了能够复性成天然态蛋白溶菌酶分子外,还会形成可溶的蛋白溶菌酶分子二聚体和三聚体,二聚体和三聚体主要是靠分子间二硫键的错配连接而成的;可溶的蛋白溶菌酶分子二聚体之间通过非共价键相互作用而形成集聚体沉淀,而可溶的三聚体溶菌酶分子则仍处于复性液上清液中。

Studies on the Aggregate During the Refolding Procedure of Reduced-denatured Egg White Lysozymes by Protein Electrophoreses and Size-exclusion Chromatography

The aggregate of egg white lysozyme molecules formed during the refolding procedure of denatured-reduced egg white lysozymes was analyzed by protein electrophoreses and high-performance size-exclusion chromatography.When the denatured-reduced egg white lysozymes were renatured by dilution method,an observable amount of aggregate precipitate could be immediately formed.The results of lauryl sodium sulfate-polyacrylamide gel electrophoreses(SDS-PAGE) of the aggregate precipitate and supernatant and the result of high-performance size-exclusion chromatography of the supernatant indicated that,by wrongly linked intermolecular disulfide bonds soluble bi-molecular and tri-molecular egg white lysozyme aggregate could be simultaneously formed except being renatured to native and active egg white lysozymes during the refolding procedure of denatured-reduced egg white lysozyme;the aggregate precipitate could be further formed by the non-covalent bonds interaction between the soluble bi-molecular egg white lysozyme aggregates,and the soluble tri-molecular egg white lysozyme aggregate could still stay at the supernatant.