Kumar Navneet, Garg Prabha
Department of Pharmacoinformatics, National Institute of Pharmaceutical Education and Research (NIPER), Sector 67, S.A.S. Nagar 160062, Punjab, India.
J Phys Chem B. 2022 Feb 24;126(7):1447-1461. doi: 10.1021/acs.jpcb.1c09438. Epub 2022 Feb 15.
The emergence of multidrug-resistant and extensively drug-resistant tuberculosis strains is the reason that the infectious tuberculosis pathogen is still the most common cause of death. The quest for new antitubercular drugs that can fit into multidrug regimens, function swiftly, and overcome the ever-increasing prevalence of drug resistance continues. The crucial role of EF-Tu in translation and trans-translation processes makes it an excellent target for antitubercular drug design. In this study, the primary sequence of EF-Tu was used to model the three-dimensional structures of EF-Tu in the presence of GDP ("off" state) and GTP ("on" state). The binding free energy computed using both the molecular mechanics/Poisson-Boltzmann surface area and umbrella sampling approaches shows that GDP binds to EF-Tu with an ∼2-fold affinity compared to GTP. The steered molecular dynamics (SMD) and umbrella sampling simulation also shows that the dissociation of GDP from EF-Tu in the presence of Mg is a thermodynamically intensive process, while in the absence of Mg, the destabilized GDP dissociates very easily from the EF-Tu. Naturally, the dissociation of Mg from the EF-Tu is facilitated by the nucleotide exchange factor EF-Ts, and this prior release of magnesium makes the dissociation process of destabilized GDP easy, similar to that observed in the umbrella sampling and SMD study. The MD simulations of EF-Tu's "on" state conformation in the presence of GTP reveal that the secondary structure of switch-I and Mg coordination network remains similar to its template despite the absence of identity in the conserved region of switch-I. On the other hand, the secondary structure in the conserved region of the switch-I of EF-Tu unwinds from a helix to a loop in the presence of GDP. The major conformational changes observed in switch-I and the movement of Thr64 away from Mg mainly reflect essential conformational changes to make the shift of EF-Tu's "on" state to the "off" state in the presence of GDP. These obtained structural and functional insights into EF-Tu are pivotal for a better understanding of structural-functional linkages of EF-Tu, and these findings may serve as a basis for the design and development of EF-Tu-specific inhibitors.
多重耐药和广泛耐药结核菌株的出现，是传染性结核病原体仍是最常见死因的原因。寻找能够纳入多药治疗方案、起效迅速且能克服日益普遍的耐药性的新型抗结核药物的工作仍在继续。EF-Tu在翻译和反式翻译过程中的关键作用，使其成为抗结核药物设计的理想靶点。在本研究中，利用EF-Tu的一级序列对其在存在GDP（“关闭”状态）和GTP（“开启”状态）时的三维结构进行建模。使用分子力学/泊松-玻尔兹曼表面积法和伞形采样法计算得到的结合自由能表明，与GTP相比，GDP与EF-Tu的结合亲和力约低2倍。引导分子动力学（SMD）和伞形采样模拟还表明，在存在Mg的情况下，GDP从EF-Tu上解离是一个热力学上的密集过程，而在不存在Mg的情况下，不稳定的GDP很容易从EF-Tu上解离。自然地，核苷酸交换因子EF-Ts促进了Mg从EF-Tu上的解离，而这种镁的预先释放使得不稳定的GDP的解离过程变得容易，这与伞形采样和SMD研究中观察到的情况类似。在存在GTP的情况下对EF-Tu“开启”状态构象的分子动力学模拟表明，尽管开关-I的保守区域缺乏一致性，但其开关-I的二级结构和Mg配位网络仍与其模板相似。另一方面，在存在GDP的情况下，EF-Tu开关-I保守区域的二级结构从螺旋展开为环。在开关-I中观察到的主要构象变化以及Thr64远离Mg的移动，主要反映了在存在GDP的情况下使EF-Tu从“开启”状态转变为“关闭”状态的基本构象变化。这些获得的关于EF-Tu的结构和功能见解对于更好地理解EF-Tu的结构-功能联系至关重要，这些发现可能为设计和开发EF-Tu特异性抑制剂提供基础。
