细胞色素b5突变体V45H NMR初步研究  

NMR Preliminary Study on Cytochrome b5 Mutant V45H

作　　者：曹春阳[1] 张乃霞[1] 王跃丰[1] 吴厚铭[1] 王志强[2] 王韵华[2] 黄仲贤[2] 

机构地区：[1]中国科学院上海有机化学研究所, [2]复旦大学化学系,

出　　处：《化学学报》2001年第4期578-586,共9页Acta Chimica Sinica

摘　　要：通过分析在H2O和D2O中采集的DQF-COSY,TOCSY和NOESY等二维核磁共振波谱鉴定了细胞色素b5定点突变体V45H (残基Val45突变为His45)的大多数氨基酸残基的质子自旋系统,通过解析NOESY谱中的dNN (i,i+1), dαN (i,i+1),dαN(i,i+2),dαN(i,i+3),dαβ(i,i+3)和dβN(i,i+1)等NOE相关,完成了其序列特异性归属以及主链和侧链质子共振信号的全归属.突变体V45H的二级结构分析表明残基Val45突变为His45对分子的整体折叠影响不大.但是,与野生型细胞色素b5相比较,突变体V45H主链酰胺质子的化学位移指数提示突变使其血红素疏水腔的微环境受到扰动.以上实验结果为进一步测定V45H的溶液结构和分析残基Val45在蛋白质中的作用提供了基础.In this paper, most of spin systems for amino acid residues in V45H, a site - directed mutant of cytochrome b5 where the residue Val(45) is mutated to His(45), have been identified through analysis of 2D NMR H-1-H-1 DQF - COSY, TOCSY and NOESY spectra acquired in H2O and D2O. The sequence - specific assignment of spin systems is obtained by d(NN)( i , i + 1), d(alphaN)( i, i + 1), d(alphaN)( i, i + 2), d(alphaN)( i, i + 3), d(alpha beta) ( i, i + 3) and d(betaN)( i, i + I) NOEs correlation found in NOESY spectra and the complete assignment of proton resonances for the backbone and side chain have been achieved. The analysis of secondary structure of V45H reveals that the mutation from Val(45) to His(45) has little effect in the global folding of the protein. However, the chemical shift index of amide protons of V45H in comparison to those of wild type cytochrome b5 indicates that the heme pocket environment is disturbed by the mutation. These experimental results provide a basis for further determination of the solution structure of V45H and the roles of residue Val(45) in the protein.

关 键 词：细胞色素B5 突变体 核磁共振 V45H 整体折叠 血红素 

分 类 号：Q559.9[生物学—生物化学]