对中国人群中III/2a型HCV高变区1序列变异规律的横断研究

Cross study of sequence diversity in genotype III/2a hepatitis C virus hypervariable region 1 of 38 Chinese patients

目的研究中国丙型肝炎患者III/2a型丙型肝炎病毒(HCV)包膜蛋白E2/NS1高变区1(HVR1)序列变异的规律及意义。方法应用逆转录巢式PCR技术,从38例III/2a型HCV感染患者血清中 ,扩增HCV部分包膜区基因片断(nt1460～1582,HCV J6),纯化后直接采用双脱氧链末端终止法进行序列分析。结果本组III/2a型HCVHVR1位于氨基酸(aa)384～408位,与有关文献报道的结果(383～410或414位)略有差异。与HCV J ,河北株、HCV 1和HCV J6相应序列比较 ,核苷酸(nt)的同源性依次为 :32 %～56 %(平均45.4 %) ,41.7 %～60.0 %(50.7 %) ,41.3 %～62.7 %(53.3 %)和49.3 %～73.3 %(58.9 %) ;aa的同源性依次为 :16 %～48 %(33.5 %) ,28 %～60 %(42.5 %) ,24 %～64 %(45.3 %)和20 %～64 %(39.4 %)。我们在本组HVR1内发现 ,5个较保守的aa位点:385位为Thr,389 ,390和406位为Gly,403位为Phe,其中390位未见于其它报道。结论本组序列变异集中于aa384～408位 ,且以异义替换为主 ,构成了免疫逃逸的基础 ,但某些位点上序列相对保守,这些位点可能是HCV抗原表位的结构位点。对HVR1序列变异规律及其生物学意义的进一步研究 ,将有助于了解HCV的致病机制及疫苗的研制。

Aim To study the sequence diversity of the hypervariable region 1(HVR1) in the putative envelope protein E2/NS1 of genotype III/2a HCV in Chinese patients. Methods The RNA(nt1460 1582,HCV J6)extracted from plasma of 38 patients infected with genotype III/2a HCV were transcribed,amplified,purified and directly sequenced by using RT nested polymerase chain reaction(PCR) and dideoxynucleotide chain termination method,respectively. Results The HVR1 was found in aa 384 408 positions of E2 protein, which was different from that in other reports. Compared with HCV J,HCV HB,HCV 1 and HCV J6, the nucleotide sequence homological rates of these HVR1 were 32% 56%(45.4%),41.7% 60.0%(50.7%),41.3% 62.7%(53.3%),49.3% 73.3%(58.9%)respectively while of homological rates of the amino acid sequence were 16% 48%(33.5%),28% 60%(42.5%),24% 64%(45.3%),20% 64%(39.4%)respectively. Five conserved residues were found in the HVR1: Thr was at position 385,Gly was at position 389,390 and 406, and Phe was at position 403, in which position 390 was not mentioned in other reports. Conclusion Our data indicated that the sequence alterations were concentrated on aa 384 408 and were mostly nonsynonymous mutations, which could help HCV to escape from host′s immune clearance. But some conserved positions were also found in these HVR1, which might provide a framework for HVR1 epitopes. Further study on the diversity of HVR1 and its biological significance will be helpful to the understanding of the mechanism of HCV persistent infection and the development of HCV vaccine.