Computational design of a thermostable mutant of cocaine esterase via molecular dynamics simulations.
Computational design of a thermostable mutant of cocaine esterase via molecular dynamics simulations.
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DOI：
10.1039/c0ob00972e
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发表时间：
2011-06-07
影响因子：
3.2
通讯作者：
Zhan CG
中科院分区：
文献类型：
--
作者：
Huang X;Gao D;Zhan CG
可卡因酯酶(Coce)被认为是代谢天然可卡因的最有效的天然酶。天然CoCE在生理温度(37℃)下的半衰期仅为~11min，其热不稳定性是其临床应用的主要障碍。开发一种耐热的Coce突变体用于治疗可卡因过量和成瘾是非常必要的。为了建立结构与热稳定性的关系，我们在400K对野生型CoCE和已知的耐热突变体进行了分子动力学(MD)模拟，表明该酶活性形式的热稳定性与模拟酶中催化残基(Y44、S117、Y118、H287和D259)的波动(表征为原子位置的RMSD和RMSF)有关。根据结构与热稳定性的相关性，进一步的计算建模，包括400K下的MD模拟，预测L169K突变体的活性中心结构应该更具热稳定性。湿实验表明，L169K突变体的活性形式在37℃下的半衰期为570min，明显长于野生型和先前已知的耐热突变体。这一令人鼓舞的结果表明，高温分子动力学模拟和结构热稳定性可能被认为是计算设计酶的耐热突变体的有价值的工具。
Cocaine esterase (CocE) has been known as the most efficient native enzyme for metabolizing the naturally occurring cocaine. A major obstacle to the clinical application of CocE is the thermoinstability of native CocE with a half-life of only ~11 min at physiological temperature (37°C). It is highly desirable to develop a thermostable mutant of CocE for therapeutic treatment of cocaine overdose and addiction. To establish a structure-thermostability relationship, we carried out molecular dynamics (MD) simulations at 400 K on wild-type CocE and previously known thermostable mutants, demonstrating that the thermostability of the active form of the enzyme correlates with the fluctuation (characterized as the RMSD and RMSF of atomic positions) of the catalytic residues (Y44, S117, Y118, H287, and D259) in the simulated enzyme. In light of the structure-thermostability correlation, further computational modeling including MD simulations at 400 K predicted that the active site structure of the L169K mutant should be more thermostable. The prediction has been confirmed by wet experimental tests showing that the active form of the L169K mutant had a half-life of 570 min at 37°C, which is significantly longer than those of the wild-type and previously known thermostable mutants. The encouraging outcome suggests that the high-temperature MD simulations and the structure-thermostability may be considered as a valuable tool for computational design of thermostable mutants of an enzyme.
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影响因子：
56.9
作者：
Korkegian, A;Black, ME;Stoddard, BL
通讯作者：
Stoddard, BL
DOI：
10.1073/pnas.0808220106
发表时间：
2009-02-24
影响因子：
11.1
作者：
Gribenko, Alexey V.;Patel, Mayank M.;Makhatadze, George I.
通讯作者：
Makhatadze, George I.
影响因子：
3.6
作者：
Cooper, Ziva D.;Narasimhan, Diwahar;Woods, James H.
通讯作者：
Woods, James H.
影响因子：
4.4
作者：
JORGENSEN, WL;CHANDRASEKHAR, J;KLEIN, ML
通讯作者：
KLEIN, ML
DOI：
10.1038/nsb742
发表时间：
2002-01-01
期刊：
NATURE STRUCTURAL BIOLOGY
影响因子：
--
作者：
Larsen, NA;Turner, JM;Wilson, IA
通讯作者：
Wilson, IA