Abstract：
SARS-CoV-2 main protease (Mpro) is a well-recognized target for COVID-19 therapy. Green tea (-)-epigallocatechin-3-gallate (EGCG) possesses Mpro-inhibitory activity; however, the influence of EGCG oxidation on its inhibition activity remains obscure, given its high oxidation propensity. This study reveals that prolonged EGCG oxidation in the presence of Mpro dramatically increases its inhibitory activity with an IC50 of 0.26 μM. The inhibitory mechanism is that EGCG-quinone preferentially binds the active site Mpro-Cys145-SH, which forms a quinoprotein. Though Mpro is present in the cell lysate, EGCG preferentially depletes its thiols. Non-cytotoxic EGCG effectively generates a quinoprotein in living cells, thus EGCG might selectively inhibit Mpro in SARS-CoV-2 infected cells. Chlorogenic acid facilitates EGCG oxidation. Together, they synergistically deplete multiple Mpro thiols though this is not more beneficial than EGCG alone. By contrast, excessive EGCG oxidation prior to incubation with Mpro largely compromises its inhibitory activity. Overall, the low IC50 and the high selectivity imply that EGCG is a promising dietary Mpro inhibitor. While EGCG oxidation in the presence of Mpro has a pivotal role in inhibition, enhancing EGCG oxidation by chlorogenic acid no longer increases its inhibitory potential. EGCG oxidation in the absence of Mpro should be avoided to maximize its Mpro-inhibitory activity.
摘要：
SARS-CoV-2主要蛋白酶（Mpro）是COVID-19治疗的公认靶标。绿茶（-）-表没食子儿茶素-3-没食子酸酯（EGCG）具有Mpro抑制活性；但是，EGCG氧化对其抑制活性的影响仍然不清楚，鉴于其高氧化倾向。该研究揭示了在Mpro存在下延长的EGCG氧化显著增加了其抑制活性,IC50为0.26μM。抑制机制是EGCG-醌优先结合活性位点Mpro-Cys145-SH，形成一种醌蛋白。虽然Mpro存在于细胞裂解物中,EGCG优先消耗其硫醇。非细胞毒性EGCG有效地在活细胞中产生一种喹蛋白，因此，EGCG可能选择性抑制SARS-CoV-2感染细胞中的Mpro。绿原酸促进EGCG氧化。一起,它们协同消耗多种Mpro硫醇，尽管这并不比单独使用EGCG更有益。相比之下,在与Mpro孵育之前过度的EGCG氧化在很大程度上损害了其抑制活性。总的来说,低IC50和高选择性意味着EGCG是一种有前途的饮食Mpro抑制剂。虽然在Mpro的存在下EGCG氧化在抑制中具有关键作用，绿原酸增强EGCG氧化不再增加其抑制潜力。应避免在不存在Mpro的情况下的EGCG氧化以使其Mpro抑制活性最大化。
