Characterization of the cofactor-induced folding mechanism of a zinc-binding peptide using computationally designed mutants.
Characterization of the cofactor-induced folding mechanism of a zinc-binding peptide using computationally designed mutants.
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DOI：
10.1016/j.jmb.2009.03.074
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发表时间：
2009-05-29
影响因子：
5.6
通讯作者：
Gai, Feng
中科院分区：
文献类型：
--
作者：
Tang, Jia;Kang, Seung-Gu;Saven, Jeffery G.;Gai, Feng
关键词：
金属是最常见的蛋白质辅因子，它们在生物学中起着重要的结构和功能作用。在许多情况下，金属结合为多肽链折叠提供主要驱动力。虽然有很多关于金属结合蛋白的结构、稳定性和功能的研究，但很少有研究关注金属诱导折叠的动力学机制。在此，研究了Zn 2+诱导的小锌结合蛋白的折叠动力学; CH 11肽衍生自转录辅助调节因子CBP的蛋白质相互作用结构域的第一个富含半胱氨酸/组氨酸的区域（CH 1结构域）。计算设计用于引入结构上与CH 11一致的色氨酸和组氨酸突变;使用停流色氨酸荧光实验研究这些突变体。Zn 2+诱导的CH 11折叠动力学与两个平行的途径一致，其中Zn 2+的初始结合发生在两个位点。然而，最初形成的Zn 2+结合的复合物可以直接进行到折叠状态，其中锌采用四面体配位或关闭路径，错连中间体。虽然消除那些负责错连接的配体简化了折叠动力学，但它也导致锌结合常数降低。因此，这些结果表明，为什么这些非天然的锌配体在CH 11结构域中是保守的几个远亲的生物体和功能的要求可能会导致动力学挫折折叠。此外，环闭合速率的CH 11肽的基础上提出的模型和温度依赖性动力学测量。
Metals are the most commonly encountered protein cofactors, and they play important structural and functional roles in biology. In many cases, metal-binding provides a major driving force for a polypeptide chain to fold. While there are many studies on the structure, stability, and function of metal-binding proteins, there are few studies focusing on the understanding of the kinetic mechanism of metal-induced folding. Herein, the Zn2+-induced folding kinetics of a small zinc-binding protein are studied; the CH11 peptide is derived from the first cysteine/histidine-rich region (CH1 domain) of protein interaction domains of the transcriptional coregulator CBP. Computational design is used to introduce tryptophan and histidine mutations that are structurally consistent with CH11; these mutants are studied using stopped-flow tryptophan fluorescence experiments. The Zn2+-induced CH11 folding kinetics are consistent with two parallel pathways, where the initial binding of Zn2+ occurs at two sites. However, the initially formed Zn2+-bound complexes can proceed either directly to the folded state where zinc adopts a tetrahedral coordination or to an off-pathway, misligated intermediate. While elimination of those ligands responsible for misligation simplifies the folding kinetics, it also leads to a decrease in the zinc binding constant. Therefore, these results suggest why these non-native zinc ligands in the CH11 domain are conserved in several distantly related organisms and that the requirement for function can lead to kinetic frustration in folding. In addition, the loop closure rate of the CH11 peptide is determined based on the proposed model and temperature dependent kinetic measurements.
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DOI：
10.1073/pnas.0709915104
发表时间：
2007-12-11
影响因子：
11.1
作者：
Ferreiro, Diego U.;Hegler, Joseph A.;Wolynes, Peter G.
通讯作者：
Wolynes, Peter G.
影响因子：
3.4
作者：
Bombarda, Elisa;Grell, Ernst;Mely, Yves
通讯作者：
Mely, Yves
影响因子：
5.6
作者：
Calhoun, JR;Kono, H;Saven, JG
通讯作者：
Saven, JG
影响因子：
3
作者：
Beck, David A. C.;White, George W. N.;Daggett, Valerie
通讯作者：
Daggett, Valerie
影响因子：
2.9
作者：
Blasie, CA;Berg, JM
通讯作者：
Berg, JM