藻蓝蛋白裂合酶CpeS色氨酸定点诱变及体内重组功能研究

Site-directed Mutagenesis of the CpeS of Phycocyanin and Study on the Function with Reconstitution in vivo

摘要: 为了研究藻蓝蛋白β亚基Cys-84裂合酶CpeS结构与功能的关系以及色氨酸残基对于该酶功能的影响,构建了藻蓝蛋白β亚基Cys-84裂合酶CpeS的两个色氨酸突变体,分别为CpeS(W14I)和CpeS(W75S)。通过体内重组检测酶活性的变化,研究色氨酸残基的突变对裂合酶催化活性的影响。重组结果显示:突变体CpeS(W14I)的催化活性几乎完全丧失,为野生型的8%;突变体CpeS(W75S)的催化活性为野生型的76%。由此推测,第14位色氨酸可能是CpeS酶活性的必需氨基酸,其所处的位置可能是裂合酶CpeS的活性位点。

Abstract: To study the structure-function relationships and the effect of the tryptophan residues to the CpeS of phycocyanin,two mutants,CpeS(W14I) and CpeS(W75S),were constructed.The lyase activity of these mutants were tested by reconstitution in vivo,to study the effects of tryptophans to CpeS.The results shows that:CpeS(W14I) has hardly lyase activity.Comparing with the wild CpeS,the lyase activity of CpeS(W14I) and CpeS(W75S) is respectively 8% and 76%.From this,it’s deduced that:the 14th tryptophan residue in CpeS may be essential amino acid to CpeS,and it may locate in the active site of CpeS.