Fisher Timothy S, Joshi Pheroze, Prasad Vinayaka R
Department of Microbiology and Immunology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
J Virol. 2002 Apr;76(8):4068-72. doi: 10.1128/jvi.76.8.4068-4072.2002.
We isolated two template analog reverse transcriptase (RT) inhibitor-resistant mutants of human immunodeficiency virus (HIV) type 1 RT by using the DNA aptamer, RT1t49. The mutations associated, N255D or N265D, displayed low-level resistance to RT1t49, while high-level resistance could be observed when both mutations were present (Dbl). Molecular clones of HIV that contained the mutations produced replication-defective virions. All three RT mutants displayed severe processivity defects. Thus, while biochemical resistance to the DNA aptamer RT1t49 can be generated in vitro via multiple mutations, the overlap between the aptamer- and template-primer-binding pockets favors mutations that also affect the RT-template-primer interaction. Therefore, viruses with such mutations are replication defective. Potent inhibition and a built-in mechanism to render aptamer-resistant viruses replication defective make this an attractive class of inhibitors.
我们使用DNA适配体RT1t49分离出了两株对模板类似物逆转录酶（RT）抑制剂耐药的1型人类免疫缺陷病毒（HIV）RT突变体。与之相关的突变N255D或N265D对RT1t49表现出低水平耐药，而当两个突变同时存在时（双突变体）则可观察到高水平耐药。含有这些突变的HIV分子克隆产生复制缺陷型病毒粒子。所有三个RT突变体均表现出严重的持续合成能力缺陷。因此，虽然通过多个突变可在体外产生对DNA适配体RT1t49的生化耐药性，但适配体与模板引物结合口袋之间的重叠有利于那些也会影响RT-模板引物相互作用的突变。所以，具有此类突变的病毒复制存在缺陷。强效抑制以及使适配体耐药病毒复制缺陷的内在机制，使得这类抑制剂颇具吸引力。
