Amino Acid Insertions near Gag Cleavage Sites Restore the Otherwise Compromised Replication of Human Immunodeficiency Virus Type 1 Variants Resistant to Protease Inhibitors
Amino Acid Insertions near Gag Cleavage Sites Restore the Otherwise Compromised Replication of Human Immunodeficiency Virus Type 1 Variants Resistant to Protease Inhibitors
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DOI：
10.1128/jvi.78.21.12030-12040.2004
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发表时间：
2004-11
影响因子：
5.4
通讯作者：
S. Tamiya;S. Mardy;M. Kavlick;K. Yoshimura;Hiroaki Mistuya
中科院分区：
文献类型：
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作者：
S. Tamiya;S. Mardy;M. Kavlick;K. Yoshimura;Hiroaki Mistuya
摘要蛋白水解酶和Gag蛋白中的多种氨基酸替换与人类免疫缺陷病毒1型(HIV-1)对蛋白水解酶抑制剂的耐药性有关。在本研究中，通过使用从有大量药物经验和治疗失败的艾滋病患者中分离出的HIV-1变异体，产生了全长的分子感染性HIV-1克隆。在产生的6个全长感染性克隆中，发现4个在p17/p24和p1/p6 Gag裂解位点附近有独特的插入(TGNS、SQVN、AQQA、SRPE、APP和/或PTAPPA)，以及已知的与抗药性相关的多个氨基酸替换。添加这样的Gag插入物主要损害了野生型HIV-1的复制，而含有插入物的原始多药耐药HIV感染克隆的复制明显好于那些修改为不含插入物的克隆。Western印迹分析表明，插入片段的存在削弱了野生型蛋白酶对Gag蛋白的加工，而突变型蛋白酶对Gag蛋白的加工能力显著提高。本研究是第一份清楚地证明在高度耐多PI的HIV-1变异体的Gag裂解位点附近看到的插入片段恢复了突变的蛋白酶的酶活性，使多PI耐药的HIV-1变异体保持复制能力的第一份报告。
ABSTRACT A variety of amino acid substitutions in the protease and Gag proteins have been reported to contribute to the development of human immunodeficiency virus type 1 (HIV-1) resistance to protease inhibitors. In the present study, full-length molecular infectious HIV-1 clones were generated by using HIV-1 variants isolated from heavily drug-experienced and therapy-failed AIDS patients. Of six full-length infectious clones generated, four were found to have unique insertions (TGNS, SQVN, AQQA, SRPE, APP, and/or PTAPPA) near the p17/p24 and p1/p6 Gag cleavage sites, in addition to the known resistance-related multiple amino acid substitutions within the protease. The addition of such Gag inserts mostly compromised the replication of wild-type HIV-1, whereas the primary multidrug-resistant HIV infectious clones containing inserts replicated significantly better than those modified to lack the inserts. Western blot analyses revealed that the processing of Gag proteins by wild-type protease was impaired by the presence of the inserts, whereas that by mutant protease was substantially improved. The present study represents the first report clearly demonstrating that the inserts seen in the proximity of the Gag cleavage sites in highly multi-PI resistant HIV-1 variants restore the otherwise compromised enzymatic activity of mutant protease, enabling the multi-PI-resistant HIV-1 variants to remain replication competent.