A Novel Regioselective Approach to Cyclize Phage-Displayed Peptides in Combination with Epitope-Directed Selection to Identify a Potent Neutralizing Macrocyclic Peptide for SARS-CoV-2
A Novel Regioselective Approach to Cyclize Phage-Displayed Peptides in Combination with Epitope-Directed Selection to Identify a Potent Neutralizing Macrocyclic Peptide for SARS-CoV-2
复制标题
结合表位定向选择环化噬菌体展示肽的新区域选择性方法，以鉴定针对 SARS-CoV-2 的有效中和大环肽
DOI：
10.1101/2022.07.06.498864
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发表时间：
2022
期刊：
影响因子：
--
通讯作者：
J. Trae Hampton, Tyler J.
中科院分区：
文献类型：
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作者：
J. Trae Hampton, Tyler J.
利用与 N 末端半胱氨酸的区域选择性氰基苯并噻唑缩合反应以及与内部半胱氨酸的氯乙酰胺反应，构建了噬菌体展示的大环 12 聚体肽库并随后进行了验证。使用该文库，结合针对新型严重急性呼吸综合征病毒 2 (SARS-CoV-2) 刺突蛋白受体结合域 (RBD) 的两个表位的迭代选择，鉴定出强烈抑制刺突 RBD 与血管紧张素转换酶 2 (ACE2)（SARS-CoV-2 人类宿主受体）之间相互作用的大环肽。使用这两个表位代替 Spike RBD，以避免选择结合 RBD 但不直接抑制其与 ACE2 相互作用的非生产性大环肽。针对 SARS-CoV-2 的抗病毒测试表明，一种大环肽对 Vero E6 细胞中的病毒繁殖具有高效能，EC50 值为 3.1 μM。 AlphaLISA 检测到的该大环肽的 IC50 值为 0.3 μM。目前的研究表明，分别针对N端和内部半胱氨酸的两个动力学控制反应在构建噬菌体展示的大环肽方面非常有效，并且基于SARS-CoV-2刺突表位的选择是鉴定肽基抗病毒药物的一种有前途的方法。
Using the regioselective cyanobenzothiazole condensation reaction with an N-terminal cysteine and the chloroacetamide reaction with an internal cysteine, a phage-displayed macrocyclic 12-mer peptide library was constructed and subsequently validated. Using this library in combination with iterative selections against two epitopes from the receptor binding domain (RBD) of the novel severe acute respiratory syndrome virus 2 (SARS-CoV-2) Spike protein, macrocyclic peptides that strongly inhibit the interaction between the Spike RBD and angiotensin-converting enzyme 2 (ACE2), the human host receptor of SARS-CoV-2, were identified. The two epitopes were used instead of the Spike RBD to avoid selection of nonproductive macrocyclic peptides that bind RBD but do not directly inhibit its interactions with ACE2. Antiviral tests against SARS-CoV-2 showed that one macrocyclic peptide is highly potent against viral reproduction in Vero E6 cells with an EC50value of 3.1 μM. The AlphaLISA-detected IC50value for this macrocyclic peptide was 0.3 μM. The current study demonstrates that two kinetically controlled reactions toward N-terminal and internal cysteines, respectively, are highly effective in the construction of phage-displayed macrocyclic peptides, and the selection based on the SARS-CoV-2 Spike epitopes is a promising methodology in the identification of peptidyl antivirals.