Patel P H, Preston B D
Laboratory for Cancer Research, Rutgers University College of Pharmacy, Piscataway, NJ 08855-0789.
Proc Natl Acad Sci U S A. 1994 Jan 18;91(2):549-53. doi: 10.1073/pnas.91.2.549.
Human immunodeficiency virus type 1 (HIV-1) is genetically highly variable. This is attributed to the error-prone nature of HIV-1 replication and its proclivity for recombination. During replication and recombination, reverse transcriptase (RT) must polymerize DNA to the 5' ends of multiple RNA and DNA template termini while converting HIV-1 RNA to double-stranded DNA. We have determined the fidelity of HIV-1 RT in vitro during polymerization to the 5' ends of HIV-1 long terminal repeat DNA template sequences and to the end of a partial HIV-1 genomic RNA template that mimics a recombination intermediate. HIV-1 RT readily extended recessed DNA primers to form full-length blunt-end DNA-DNA and DNA-RNA duplexes. In addition, HIV-1 RT catalyzed high yields of products with one to four extra nucleotides at the 3' ends of the nascent DNAs. These products were formed processively via a nontemplated mechanism that is highly specific for the addition of purine nucleotides (A > G >> T > or = C). Thus, HIV-1 RT is extremely unfaithful at both DNA and RNA template ends, introducing errors (extra nucleotides) in one out of every two or three nascent strands processively polymerized. This error rate is 1000 times higher than for HIV-1 RT-catalyzed errors at internal template positions. Blunt-end additions were also catalyzed by other retroviral RTs at relative rates of HIV-1 approximately Moloney murine leukemia virus > avian myeloblastosis virus. These data suggest a potentially important mechanism for retroviral mutation mediated by nontemplated blunt-end addition of purines prior to forced copy-choice recombination.
1型人类免疫缺陷病毒（HIV-1）在基因上具有高度变异性。这归因于HIV-1复制的易错性及其重组倾向。在复制和重组过程中，逆转录酶（RT）必须将DNA聚合到多个RNA和DNA模板末端的5'端，同时将HIV-1 RNA转化为双链DNA。我们已经确定了HIV-1 RT在体外将DNA聚合到HIV-1长末端重复DNA模板序列的5'端以及模拟重组中间体的部分HIV-1基因组RNA模板末端时的保真度。HIV-1 RT很容易延伸凹陷的DNA引物，以形成全长平端DNA-DNA和DNA-RNA双链体。此外，HIV-1 RT催化在新生DNA的3'端产生一到四个额外核苷酸的高产率产物。这些产物通过一种非模板机制连续形成，该机制对嘌呤核苷酸的添加具有高度特异性（A>G>>T>或=C）。因此，HIV-1 RT在DNA和RNA模板末端都极其不忠实，在每两到三条连续聚合的新生链中就有一条引入错误（额外核苷酸）。这个错误率比HIV-1 RT在内部模板位置催化的错误率高1000倍。其他逆转录病毒RT也以大约HIV-1>莫洛尼鼠白血病病毒>禽成髓细胞瘤病毒的相对速率催化平端添加。这些数据表明，在强制拷贝选择重组之前，通过嘌呤的非模板平端添加介导逆转录病毒突变的潜在重要机制。
