Gene expression of NS5a strong antigenic regions of hepatitis C virus in E. coli and detection of their antigenicity

作     者：Howard Fields 

作者机构：the Centers for Disease Control and Prevention USA 

出 版 物：《Hepatobiliary & Pancreatic Diseases International》 (国际肝胆胰疾病杂志（英文版）)

年 卷 期：2002年第1卷第3期

页      面：383-387页

学科分类：1001[医学-基础医学(可授医学、理学学位)] 100102[医学-免疫学] 10[医学] 

主　　题：hepatitis C virus gene expression anti genie heterogeneity 

摘      要：Objective: To study the effect of sequence variability between different genotypes of HCV on the antigenic properties of the NS5a protein of strong antigenic re- gion at position 2212-2313 by using recombinant proteins. Methods: Thirteen representative sequences from HCV genotypes 1 to 6 were selected to design synthe- tic genes encoding for this antigenic region, These genes were assembled by PCR from synthetic olionu- cleotides and expressed in E. coli as hybrid proteins with glutathione S-transferase. All 13 fusion proteins were purified from bacterial lysates and used to test a panel of anti-HCV positive sera (n=61) obtained from patients infected with HCV genotypes 1 through 6. Results: A comparison of sequences derived from dif- ferent HCV genotypes showed that the primary struc- ture of this strong antigenic region is highly variable. Percent homology between different genotype se- quences varied from 40. 4% to 72. 5%. All but one protein immunoreacted with 62% to 93% of serum samples. Although a variable degree of genotype spe- cific antigenic reactivity was detected, only one pro- tein demonstrated a noticeable preference to immu- noreact with antibodies against the homologous HCV genotype. On the other hand, closely related proteins derived from the same subtype or genotype immuno- reacted with significantly different efficiency with HCV antibodies. Conclusions: Different genotype HCV genes were successfully cloned, expressed and purified. Se- quence variability has a profound effect on the anti- genic properties of the HCV NS5a immunodominant region. This finding should be considered in the de- velopment of diagnostic tests for the efficient detec- tion of anti-HCV activity in serum specimens.