Analysis of human immunodeficiency virus type 1 reverse transcriptase subunit structure/function in the context of infectious virions and human target cells.
Analysis of human immunodeficiency virus type 1 reverse transcriptase subunit structure/function in the context of infectious virions and human target cells.
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在感染性病毒粒子和人类靶细胞的背景下分析人类免疫缺陷病毒 1 型逆转录酶亚基结构/功能。
DOI：
10.1128/aac.49.9.3762-3769.2005
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发表时间：
2005
期刊：
影响因子：
--
通讯作者：
Kappes,JohnC
中科院分区：
文献类型：
--
作者：
Mulky,Alok;Kappes,JohnC
The reverse transcriptase (RT) of all retroviruses is required for synthesis of the viral DNA genome. The human immunodeficiency virus type 1 (HIV-1) RT exists as a heterodimer made up of 51-kDa and 66-kDa subunits. The crystal structure and in vitro biochemical analyses indicate that the p66 subunit of RT is primarily responsible for the enzyme's polymerase and RNase H activities. Since both the p51 and p66 subunits are generated from the same coding region, as part of the Pr160Gag-Polprecursor protein, there are inherent limitations for studying subunit-specific function with intact provirus in a virologically relevant context. Our lab has recently described a novel system for studying the RT heterodimer (p51/p66) wherein a LTR-vpr-p51-IRES-p66 expression cassette provided intransto an RT-deleted HIV-1 genome allows precise molecular analysis of the RT heterodimer. In this report, we describe in detail the specific approaches, alternative strategies, and pitfalls that may affect the application of this novel assay for analyzing RT subunit structure/function in infectious virions and human target cells. The ability to study HIV-1 RT subunit structure/function in a physiologically relevant context will advance our understanding of both RT and the process of reverse transcription. The study of antiretroviral drugs in a subunit-specific virologic context should provide new insights into drug resistance and viral fitness. Finally, we anticipate that this approach will help elucidate determinants that mediate p51-p66 subunit interactions, which is essential for structure-based drug design targeting RT heterodimerization.
