摘要
将野生型鸡心脱辅基细胞色素c及其突变体V92A的17位半胱氨酸残基突变为丝氨酸,再将表达纯化的V92A/C17S和W/C17S用荧光探针IAEDANS标记.通过测量AEDANS-Cys-14的荧光光谱、荧光寿命以及AEDANS与Trp-59之间的荧光共振能量转移效率、比较了V92A与野生型鸡心脱辅基细胞色素c因折叠状态不同引起的N端构象状态及肽链间相互作用的差异.结果显示Apo.c无论在低盐浓度下的无规卷曲状态还是高盐浓度下的融球态,V92A都较野生型处于更松散的折叠状态,此外。
It has been reported that chicken heart apocytochrome c has been shown to possess a much stronger tendency to fold spontaneously in aqueous solution than the equivalent from other species and V92A could alter the folding propensity of chicken apocytochrome c and its interaction with phospholipid.In order to further study the conformation difference between wild type of chicken heart apocytochrome c and its V92A mutant,two mutants,W/C17S and V92A/C17S,in which the 17 cysteine residue was replaced by serine were obtained and expressed in E.coli .By using fluorescence probe IAEDANS(reacted on the SH group of 14 cystein residue) the conformation and that of intramolecular interactions between W/C17S and V92A/C17S were investigated by AEDANS fluorescence emission spectra,fluorescence lifetime and fluorescence energy transfer efficiency.Obtained results showed that comparing with the wild type of chicken heart apocytochrome c the V92A mutant appeared less folded state both in the presence and absence of 5 0 mol/L NaClO 4.It also seemed that the N terminal of the chicken heart apocytochrome c might not be involved during its spontaneous folding.
出处
《中国生物化学与分子生物学报》
CAS
CSCD
1999年第1期92-97,共6页
Chinese Journal of Biochemistry and Molecular Biology
基金
国家自然科学基金委员会重点项目
中国科学院重大项目
关键词
Apo.c
自发折叠
AEDANS标记
Apocytochrome c,Folding,AEDANS label,Fluorescence energy transfer efficiency