抗菌肽urumin与甲型流感病毒H1N1 HA蛋白的相互作用研究

Interaction study of the antibacterial peptide urumin with the H1N1 HA protein of influenza A virus

甲型H1N1流感严重影响人类的健康和破坏全球经济发展,抗菌肽urumin可特异性结合H1N1病毒血凝素(hemagglutinin, HA)蛋白的保守茎部,但其结合位点和作用机制尚不明确。本研究通过分子对接和ELISA (enzymelinked immunosorbent assay)实验研究urumin与HA蛋白作用可能的结合位点、关键氨基酸,提示HA残基His32(HA1)、Asp19 (HA2)和Trp21 (HA2)是HA与urumin作用的关键残基。Urumin的Arg4、Asn9和Cys16与HA蛋白残基Asp19 (HA2)、Trp21 (HA2)、His32 (HA1)和Asn53 (HA2)形成氢键相互作用, Trp12与HA的His32 (HA1)形成芳香π堆积作用,维持urumin与HA蛋白的结合。在293T细胞中表达野生型HA及其丙氨酸突变体[丙氨酸替换His32(HA1)、Asp19 (HA2)和Trp21 (HA2)], ELISA实验结果显示, urumin与野生型HA的亲和力显著高于HA丙氨酸突变体,提示His32 (HA1)、Asp19 (HA2)和Trp21 (HA2)可能是HA与urumin作用的关键残基。本研究为urumin的进一步改造和应用提供了理论和实验基础。

Influenza A virus(H1N1) seriously affects the health of human and disrupts the development of global economic. The antimicrobial peptide urumin specifically binds to the conserved stem of the hemagglutinin(HA) protein of H1N1 virus, but its binding site and the mechanism of action are not clear. In this study, we investigated the possible binding sites and key amino acids for the interaction of urumin with HA protein by molecular docking and enzyme-linked immunosorbent assay(ELISA) experiments, suggesting that HA residues His32(HA1), Asp19(HA2), and Trp21(HA2) are the key residues for the interaction of HA with urumin. Urumin’s Arg4,Asn9, and Cys16 were associated with HA protein residues Asp19(HA2), Trp21(HA2), His32(HA1), and Asn53(HA2) form hydrogen bonding interactions, and Trp12 forms an aromatic π-stacking interaction with His32(HA1)of HA, these interactions maintain the binding of urumin to HA protein. Wild-type HA and its alanine mutant [alanine substitutions His32(HA1), Asp19(HA2), and Trp21(HA2)] were expressed in 293 T cells. ELISA experiments showed that the affinity ability of urumin with HA wild-type was significantly higher than that of HA alanine mutant, suggesting that His32(HA1), Asp19(HA2), and Trp21(HA2) may be the key residues for HA to interact with urumin. This study provides a theoretical and experimental basis for further modification and application of urumin.