HIV-1包膜糖蛋白gp120 C3～C4区中和抗体保守表位氨基酸变异研究

Study on amino acid mutation in conserved neutralization epitopes in HIV-1 gp120 C3-C4 region

目的:研究HIV-1包膜糖蛋白gp120C3～C4区中和抗体保守表位氨基酸变异特征,探讨中和抗体表位变异与疾病进展关系,为开展中和抗体免疫治疗及疫苗设计奠定理论基础。方法:RT-PCR及nest-PCR扩增无症状HIV感染者、AIDS患者及疾病长期不进展者HIV-1gp120env C3～C4区基因,双脱氧终止法进行核酸序列测定,翻译为氨基酸序列,与HIV-1Se-quence Database参考毒株比对识别中和抗体保守表位氨基酸的变异。结果:HIVgp120C3～C4区,无症状HIV感染者/AIDS患者CD4结合位点(CD4BS)、CD4诱导(CD4i)、2G12中和抗体保守表位氨基酸均存在变异,长期不进展者2G12中和抗体表位氨基酸存在变异,三组病例表位突变率差异未达显著性(P>0.05);CD4BS、CD4i、2G12变异表位构成比分别为15.8%、31.6%、52.6%,2G12表位变异高于CD4BS和CD4i表位,差异具有显著性(P<0.05);CD4BS表位变异见于K421R,CD4i表位变异见于R419S/K,K421R,均为单位点氨基酸变异,2G12表位变异见于S334N,N332E/Q,N386D,N392S/T,N448K/I,单、双位点氨基酸变异各半。结论:HIV-1包膜糖蛋白gp120C3～C4区,无症状HIV感染者/AIDS患者CD4BS、CD4i、2G12中和抗体保守表位氨基酸均存在变异,长期不进展者表位氨基酸相对稳定,目前发现2G12表位存在变异。中和抗体表位中,2G12表位变异较CD4BS和CD4i表位多见。不同类型中和抗体保守表位氨基酸位点变异程度存在差异。

Objective： To study the mutation of amino acids in conserved neutralization epitopes in HIV-1 gp120 C3 - CA region,to discuss the relationship between the conserved neutralization epitopes and disease progression, and to provide a basis for design of vaccine and immunotherapy with neutralizing antibedy.Methods： RT-PCR and nest-PCR methods were used to amplify gene in HIV-1 gp120 C3 - C4 region. The sequence of nucleic acids was detected by double-deoxygen terminal method and translated into amino acids for analysis. Mutations of conserved neutralization epitopes was identified by comparison with the epitopes reference data in HIV-1 Sequence Database. Results： Both asymptomatic HIV patients and MDS patients had mutation in conserved neutralization epitopes, including CD4-hinding site （CD4BS）, C3）4-induced （CD4i） and 2G12 epitopes, whereas the long-term non-pmgressors （LTNPs） had 2G12 epitope mutation. The mutation rates had no significant differences （ P 〉 0.05）among HIV/AIDS patients and LTNPs. The constituent ratios of mutation epitopes of CD4BS, CD4i and 2G12 were respectively 15. 8 %, 31. 6 % and 52.6 % （ P 〈 0.05）. Mutation of amino acids in CIMBS and CD4i epitopes predominantly happened at single-site. The mutation of CD4BS epitopes was mainly K421R, whereas the mutations in CD4i epitopes were mainly R419S/K or K421R. Double-site mutation amino acids in 2G12 epitopes had a half proportion.2G12 epitope mutations were predominantly S334N, N332E/Q, N386D,N392S/T and N448K/I. Conclusion： In HIV-1 gp120 C3- CA region,there exist amino acids mutations in conserved neutralization epitopes,including CD4BS, CD4i and 2G12 epitopes, whereas the virus in LTNPs has stable conserved neutralization epitopes and rarely happens amino acids mutation except 2G12 epitopes. Among mutant epitopes, the constituent ratio for 2G12 epitope is higher than that of CD4BS and CD4i epitopes. The mutation degrees of amino acids in conserved neutralization epitopes are different.