Chemical-induced unfolding of cofactor-free protein monitored by electrochemistry
Chemical-induced unfolding of cofactor-free protein monitored by electrochemistry
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DOI：
10.1021/ac060351h
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发表时间：
2006-09-01
影响因子：
7.4
通讯作者：
Qu, Na
中科院分区：
文献类型：
--
作者：
Guo, Liang-Hong;Qu, Na
为了更好地理解蛋白质序列、结构和功能之间的关系，蛋白质折叠已经被广泛研究。已经开发并利用了大量的技术来探测蛋白质构象和折叠/解折叠转变。在这份报告中，电化学监测尿素诱导展开的一个大的无辅因子的蛋白质，牛血清白蛋白（BSA），进行了描述。以Os（bpy）（2）dppz（bpy = 2，2 '-bipyridine，dppz = dipyrido[3，2- a：2'，3 '-c] phenazine）为电子介质，在氧化铟锡电极上实现了酪氨酸和色氨酸在游离氨基酸和牛血清白蛋白中的电化学氧化。氧化电流被用作监测尿素诱导的BSA变性的信号报告。在高尿素浓度下，电化学信号相对于天然蛋白质增加了3倍。这种增加归因于未折叠BSA中的可氧化残基与Os（bpy）（2）dppz之间的更紧密接触。通过电化学评估的展开程度与在0-10 M尿素范围内建立的荧光技术良好相关。该方法可用于研究其他无辅因子蛋白质的去折叠过程。
Protein folding has been studied extensively with an aim to better understanding of the relationship between protein sequence, structure, and function. A large variety of techniques have been developed and utilized to probe protein conformation and folding/unfolding transition. In this report, electrochemical monitoring of urea-induced unfolding of a large cofactor-free protein, bovine serum albumin (BSA), is described. Enhanced electrochemical oxidation of tyrosine and tryptophan in free amino acids and in BSA was achieved on an indium tin oxide electrode by using an electron mediator, Os(bpy) (2)dppz (bpy = 2,2'-bipyridine, dppz = dipyrido[3,2- a: 2', 3'-c] phenazine). The oxidation current was used as a signal reporter in the monitoring of urea-induced BSA denaturation. At high urea concentrations, the electrochemical signal increased by 3-fold relative to the native protein. The increase is attributed to the closer contact between the oxidizable residues in the unfolded BSA and Os(bpy)(2)dppz. The degree of unfolding assessed by electrochemistry correlates well with the established fluorescence technique in the range of 0-10 M urea. The method can be used to investigate the unfolding process of other cofactor-free proteins.