病毒性趋化因子vMIP2的表达、纯化及生物活性研究

趋化因子受体如CCR5和CXCR4是HIV侵入细胞的辅助受体，趋化因子与其受体的结合可以抑制HIV感染细胞。近年来在疱疹病毒8(Human herpesvirus8，HHV8)基因组中发现与人趋化因子有较高同源性的开放阅读框，分别命名为vMIP1、vMIP2和vMIP3。研究发现vMIP2与多种人趋化因子受体有高亲和力。本研究在大肠杆菌中表达出融合蛋白TrxA—vMIP2，用亲和层析的方法对其纯化。纯化产物用肠激酶酶切后，经离子交换层析纯化出目的蛋白vMIP2。体外活性研究表明纯化的vMIP2可以有效地抑制R5和X4 HIV—1在人外周血单核细胞上的复制。

Expression, Purification and Biological Activity Study of Viral Chemokine vMIP2

Chemokine receptor CCR5 and CXCR4 are principal coreceptors for HIV. Blockage of Human immunodeficiecy virus (HIV) infection can be achieved by engaging CCR5 and CXCR4 with their natural ligands. Human herpesvirus 8 encodes three chemokines., vMIP1, vMIP2 and vMIP3, vMIP2 has been shown to bind a range of receptors including CCR5 and CXCR4. In this study, we report the expression and purification of recombinant vMIP2 from Escherichia coli. vMIP2 gene was cloned into pET-32a(+) expression vector, which allows production of the desired protein along with a thioredoxin fusion tag. The vector containing the sequence encoding the mature form of vMIP2 was transformed into AD494(DE3). After induction, TrxA- vMIP2 fusion protein was purified using Ni chelating column. Cleavage of the thioredoxin fusion tag was subsequently carried out with enterokinase. The cleaved protein was further purified by cation exchange column. Western blotting indicated that purified vMIP2 had specific immunological activity with vMIP2 antibody. In vitro infection demonstrated that vMIP2 potently inhibited the replication of R5 and X4 HIV in human peripheral blood mononuclear cell (PBMC). This study provides basis for development of effective prevention strategies against HIV. Further investigation may help to define the role of vMIP2 in HHV8 pathogenesis.