An Active Site Inhibitor Induces Conformational Penalties for ACE2 Recognition by the Spike Protein of SARS-CoV‑2

The
novel RNA virus, severe acute respiratory syndrome coronavirus
II (SARS-CoV-2), is currently the leading cause of mortality in 2020,
having led to over 1.6 million deaths and infecting over 75 million
people worldwide by December 2020. While vaccination has started and
several clinical trials for a number of vaccines are currently underway,
there is a pressing need for a cure for those already infected with
the virus. Of particular interest in the design of anti-SARS-CoV-2
therapeutics is the human protein angiotensin converting enzyme II
(ACE2) to which this virus adheres before entry into the host cell.
The SARS-CoV-2 virion binds to cell-surface bound ACE2 via interactions
of the spike protein (s-protein) on the viral surface with ACE2. In
this paper, we use all-atom molecular dynamics simulations and binding
enthalpy calculations to determine the effect that a bound ACE2 active
site inhibitor (MLN-4760) would have on the binding affinity of SARS-CoV-2
s-protein with ACE2. Our analysis indicates that the binding enthalpy
could be reduced for s-protein adherence to the active site inhibitor-bound
ACE2 protein by as much as 1.48-fold as an upper limit. This weakening
of binding strength was observed to be due to the destabilization
of the interactions between ACE2 residues Glu-35, Glu-37, Tyr-83,
Lys-353, and Arg-393 and the SARS-CoV-2 s-protein receptor binding
domain (RBD). The conformational changes were shown to lead to weakening
of ACE2 interactions with SARS-CoV-2 s-protein, therefore reducing
s-protein binding strength. Further, we observed increased conformational
lability of the N-terminal helix and a conformational shift of a significant
portion of the ACE2 motifs involved in s-protein binding, which may
affect the kinetics of the s-protein binding when the small molecule
inhibitor is bound to the ACE2 active site. These observations suggest
potential new ways for interfering with the SARS-CoV-2 adhesion by
modulating ACE2 conformation through distal active site inhibitor
binding.

新型RNA病毒严重急性呼吸综合征冠状病毒2（SARS-CoV-2）是2020年全球致死率最高的病原体，截至2020年12月，该病毒已在全球造成超160万人死亡、7500余万人感染。尽管疫苗接种工作已启动，多款疫苗的临床试验也正在推进，但目前仍亟需针对已感染患者的治疗方案。

在抗SARS-CoV-2治疗药物的研发中，人类血管紧张素转换酶2（ACE2）是备受关注的关键靶点：该病毒在侵入宿主细胞前，会通过自身表面的刺突蛋白（spike protein，简称S蛋白）与ACE2结合，从而黏附于宿主细胞表面。

本研究通过全原子分子动力学模拟与结合焓计算，探究了ACE2活性位点抑制剂MLN-4760的结合对SARS-CoV-2 S蛋白与ACE2结合亲和力的影响。分析结果显示，与结合了MLN-4760的ACE2结合时，S蛋白的结合焓最高可降低1.48倍，达到理论上限。

结合强度的减弱可归因于ACE2残基Glu-35、Glu-37、Tyr-83、Lys-353与Arg-393与SARS-CoV-2 S蛋白受体结合域（receptor binding domain，简称RBD）之间的相互作用稳定性下降。构象变化会削弱ACE2与SARS-CoV-2 S蛋白的相互作用，进而降低S蛋白的结合强度。

此外，本研究还观察到，当小分子抑制剂结合至ACE2活性位点后，ACE2的N端螺旋构象不稳定性升高，且参与S蛋白结合的多个ACE2基序发生了显著构象偏移，这可能会影响S蛋白结合的动力学过程。

上述研究结果表明，通过远端活性位点抑制剂结合来调控ACE2构象，或可成为干扰SARS-CoV-2病毒黏附的全新潜在策略。