Mello I M V G C, Thumann C, Schvoerer E, Soulier E, Pinho J R R, Silvestre D A M M, Queiroz A T L, Wolf P, Baumert T F, Keller F S, Pereira C A
Instituto Butantan, Laboratório de Imunologia Viral, São Paulo, Brazil.
J Viral Hepat. 2009 Oct;16(10):732-7. doi: 10.1111/j.1365-2893.2009.01125.x. Epub 2009 Mar 25.
As a consequence of selective pressure exerted by the immune response during hepatitis C virus (HCV) infection, a high rate of nucleotide mutations in the viral genome is observed which leads to the emergence of viral escape mutants. The aim of this study was to evaluate the evolution of the amino acid (aa) sequence of the HCV nonstructural protein 3 (NS3) in viral isolates after liver transplantation. Six patients with HCV-induced liver disease undergoing liver transplantation (LT) were followed up for sequence analysis. Hepatitis C recurrence was observed in all patients after LT. The rate of synonymous (dS) nucleotide substitutions was much higher than that of nonsynonymous (dN) ones in the NS3 encoding region. The high values of the dS/dN ratios suggest no sustained adaptive evolution selection pressure and, therefore, absence of specific NS3 viral populations. Clinical genotype assignments were supported by phylogenetic analysis. Serial samples from each patient showed lower mean nucleotide genetic distance when compared with samples of the same HCV genotype and subtype. The NS3 samples studied had an N-terminal aa sequence with several differences as compared with reference ones, mainly in genotype 1b-infected patients. After LT, as compared with the sequences before, a few reverted aa substitutions and several established aa substitutions were observed at the N-terminal of NS3. Sites described to be involved in important functions of NS3, notably those of the catalytic triad and zinc binding, remained unaltered in terms of aa sequence. Rare or frequent aa substitutions occurred indiscriminately in different positions. Several cytotoxic T lymphocyte epitopes described for HCV were present in our 1b samples. Nevertheless, the deduced secondary structure of the NS3 protease showed a few alterations in samples from genotype 3a patients, but none were seen in 1b cases. Our data, obtained from patients under important selective pressure during LT, show that the NS3 protease remains well conserved, mainly in HCV 3a patients. It reinforces its potential use as an antigenic candidate for further studies aiming at the development of a protective immune response.
由于丙型肝炎病毒（HCV）感染期间免疫反应施加的选择压力，在病毒基因组中观察到高比率的核苷酸突变，这导致病毒逃逸突变体的出现。本研究的目的是评估肝移植后病毒分离株中HCV非结构蛋白3（NS3）氨基酸（aa）序列的演变。对6例因HCV引起的肝病接受肝移植（LT）的患者进行随访以进行序列分析。LT后所有患者均观察到丙型肝炎复发。在NS3编码区，同义（dS）核苷酸替换率远高于非同义（dN）替换率。dS/dN比值的高值表明没有持续的适应性进化选择压力，因此不存在特定的NS3病毒群体。系统发育分析支持临床基因型分类。与相同HCV基因型和亚型的样本相比，每位患者的系列样本显示出较低的平均核苷酸遗传距离。与参考序列相比，所研究的NS3样本的N端氨基酸序列存在一些差异，主要见于1b基因型感染患者。LT后，与之前的序列相比，在NS3的N端观察到一些回复性氨基酸替换和几个确定的氨基酸替换。描述为参与NS3重要功能的位点，特别是催化三联体和锌结合位点，在氨基酸序列方面保持不变。罕见或频繁的氨基酸替换在不同位置随机发生。我们的1b样本中存在几种针对HCV描述的细胞毒性T淋巴细胞表位。然而，NS3蛋白酶的推导二级结构在3a基因型患者的样本中显示出一些改变，但在1b病例中未观察到。我们从LT期间处于重要选择压力下的患者获得的数据表明，NS3蛋白酶保持良好保守，主要在HCV 3a患者中。这加强了其作为抗原候选物在旨在开发保护性免疫反应的进一步研究中的潜在用途。
