Delfing Bryan M, Laracuente Xavier E, Luo Xingyu, Olson Audrey, Jeffries William, Foreman Kenneth W, Paige Mikell, Kehn-Hall Kylene, Lockhart Christopher, Klimov Dmitri K
School of Systems Biology, George Mason University, Manassas, Virginia 20110, United States.
Department of Chemistry and Biochemistry, George Mason University, Fairfax, Virginia 22030, United States.
ACS Omega. 2024 Sep 16;9(38):40259-40268. doi: 10.1021/acsomega.4c06981. eCollection 2024 Sep 24.
Several small molecule inhibitors have been designed to block binding of the Venezuelan equine encephalitis virus (VEEV) nuclear localization signal (NLS) sequence to the importin-α nuclear transport protein. To probe the inhibition mechanism on a molecular level, we used all-atom explicit water replica exchange molecular dynamics to study the binding of two inhibitors, I1 and I2, to the coreNLS peptide, representing the core fragment of the VEEV NLS sequence. Our objective was to evaluate the possibility of masking wherein binding of these inhibitors to the coreNLS occurs prior to its binding to importin-α. We found that the free energy of I1 and I2 binding to the coreNLS is less favorable than that to importin-α. This outcome argues against preemptive inhibitor binding to the coreNLS prior to importin-α. Instead, both inhibitors are expected to compete with the coreNLS peptide for binding to importin-α. The two factors responsible for the low affinities of the inhibitors to the coreNLS peptide are (i) the low cooperativity of binding to the peptide and (ii) the strong hydrophobic effect associated with binding to importin-α. Our results further show that upon binding to the coreNLS peptide, the inhibitors form multiple diverse binding poses. The coreNLS peptide coincubated with I1 and I2 adopts several conformational states, including open and collapsed, which underscores the fluidity of the coreNLS conformational ensemble as a target for inhibitors. Taken together with our prior investigations, this study sheds light on the molecular mechanism by which I1 and I2 ligands inhibit binding of the VEEV capsid protein to importin-α.
已经设计了几种小分子抑制剂来阻断委内瑞拉马脑炎病毒(VEEV)核定位信号(NLS)序列与输入蛋白-α核转运蛋白的结合。为了在分子水平上探究抑制机制,我们使用全原子显式水复制交换分子动力学来研究两种抑制剂I1和I2与coreNLS肽的结合,该肽代表VEEV NLS序列的核心片段。我们的目标是评估掩蔽的可能性,即这些抑制剂在与输入蛋白-α结合之前先与coreNLS结合。我们发现I1和I2与coreNLS结合的自由能比与输入蛋白-α结合的自由能更不利。这一结果反对抑制剂在输入蛋白-α之前抢先与coreNLS结合。相反,预计两种抑制剂都会与coreNLS肽竞争与输入蛋白-α的结合。导致抑制剂与coreNLS肽亲和力低的两个因素是:(i)与肽结合的低协同性和(ii)与输入蛋白-α结合相关的强疏水效应。我们的结果进一步表明,与coreNLS肽结合后,抑制剂形成多种不同的结合构象。与I1和I2共同孵育的coreNLS肽呈现几种构象状态,包括开放和折叠状态,这突出了coreNLS构象集合作为抑制剂靶点的流动性。结合我们之前的研究,这项研究揭示了I1和I2配体抑制VEEV衣壳蛋白与输入蛋白-α结合的分子机制。