Conformational change of the methionine 20 loop of Escherichia coli dihydrofolate reductase modulates pKa of the bound dihydrofolate
Conformational change of the methionine 20 loop of Escherichia coli dihydrofolate reductase modulates pKa of the bound dihydrofolate
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DOI：
10.1110/ps.062724307
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发表时间：
2007-06-01
期刊：
影响因子：
8
通讯作者：
Brooks, Charles L., III
中科院分区：
文献类型：
--
作者：
Khavrutskii, Ilja V.;Price, Daniel J.;Brooks, Charles L., III
通过自由能微扰和分子动力学模拟（FEP/MD），计算了大肠杆菌二氢叶酸还原酶（ecDHFR）与二氢叶酸（H2 F）形成的三种三元复合物，即封闭形式的ecDHFR（NADP（+）：H2 F）（1）、封闭形式的Michaelis复合物（2）和封闭形式的Michaelis复合物（3）中N-5位二氢叶酸（H2 F）的pK（a）。我们的模拟表明，在米氏复合物中，pK（a）是由Met 20环波动调制的，提供了最大的pK（a）位移与“紧闭”“环构象的亚态;在“部分闭合/开放”亚态中，pK（a）与封闭复合物中的pK（a）相似。有利于质子化，紧密闭合Met 20环增强了辅因子和底物与Met 20侧链的相互作用，并使辅因子的烟酰胺环与底物的蝶呤环共面排列。总体而言，本研究支持N-5直接从溶液中质子化的假设，并提供了对底物质子化机制的进一步见解。
We evaluate the pK(a) of dihydrofolate (H2F) at the N-5 position in three ternary complexes with Escherichia coli dihydrofolate reductase (ecDHFR), namely ecDHFR(NADP(+):H2F) in the closed form (1), and the Michaelis complexes ecDHFR(NADPH: H2F) in the closed (2) and occluded (3) forms, by performing free energy perturbation with molecular dynamics simulations (FEP/MD). Our simulations suggest that in the Michaelis complex the pK(a) is modulated by the Met20 loop fluctuations, providing the largest pK(a) shift in substates with a "tightly closed'' loop conformation; in the "partially closed/ open'' substates, the pK(a) is similar to that in the occluded complex. Conducive to the protonation, tightly closing the Met20 loop enhances the interactions of the cofactor and the substrate with the Met20 side chain and aligns the nicotinamide ring of the cofactor coplanar with the pterin ring of the substrate. Overall, the present study favors the hypothesis that N-5 is protonated directly from solution and provides further insights into the mechanism of the substrate protonation.