新型冠状病毒受体结合域-Fc融合蛋白表达及小鼠免疫原性

Expression and immunogenicity in mouse of SARS-CoV-2 receptor binding domain-Fc fusion protein

目的表达制备新型冠状病毒刺突蛋白的受体结合域(receptor binding domain, RBD)-Fc融合蛋白, 并评价其在小鼠模型中的免疫原性。方法将新型冠状病毒RBD与小鼠免疫球蛋白Fc段融合基因在中国仓鼠卵巢细胞中表达并制备融合蛋白。将不同剂量的RBD-Fc融合蛋白分别单独或辅以氢氧化铝佐剂免疫小鼠, 通过ELISA、假病毒中和试验、酶联免疫斑点试验检测诱导产生的体液和细胞免疫应答。结果 10 μg RBD-Fc融合蛋白辅以氢氧化铝佐剂能有效诱导小鼠产生良好的RBD特异性IgG抗体应答, 该抗体可阻断病毒RBD与细胞表面病毒受体的结合, 进一步研究表明该重组蛋白还诱导产生了针对原始毒株、Beta变异株和Delta变异株假病毒的中和抗体, 中和抗体滴度分别为1 566、336和270。结论制备的RBD-Fc融合蛋白具有较好的免疫原性, 可为开发COVID-19重组蛋白疫苗提供参考。

Objective To express the severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)spike protein receptor binding domain(RBD)-Fc fusion protein and investigate its immunogenicity in mice.Methods The fusion gene of SARS-CoV-2 RBD and Fc segment of mouse immunoglobulin was expressed in Chinese hamster ovary cells to obtain the fusion protein.Mice were immunized with different doses of RBD-Fc fusion protein alone or combined with aluminum hydroxide adjuvant.The humoral and cellular immune responses induced by RBD-Fc fusion protein were detected by ELISA,pseudovirus neutralization and enzyme-linked immunospot assay.Results Ten-microgram RBD-Fc fusion protein combined with aluminum hydroxide adjuvant effectively elicited high titers of RBD-specific IgG antibodies in immunized mice,which could block the binding of RBD to viral receptors on cell surface.Further assay showed that RBD-Fc protein induced a high level of neutralizing antibodies against the wild-type pseudovirus and a certain degree of cross-neutralizing antibodies against pseudoviruses of Beta and Delta variants(titers at 1566,336 and 270,respectively).Conclusion The RBD-Fc fusion protein has good immunogenicity,which can provide reference for the development of COVID-19 recombinant protein vaccines.