Wöhrl B M, Moelling K
Max-Planck-Institut für Molekulare Genetik, Abt. Schuster, Germany.
Behring Inst Mitt. 1991 Jul(89):100-7.
The retroviral replicating enzyme reverse transcriptase (RT) is associated with an RNase H which specifically hydrolyzes RNA in RNA-DNA hybrids. The RNase H exhibits endo- as well as exonuclease activity when analyzed with in vitro synthesized viral RNA hybridized to a shorter synthetic DNA oligonucleotide. In the presence of deoxyribonucleotides the RT synthesizes DNA in a concerted action with the RNase H, which simultaneously hydrolyzes the RNA template. Mutants of the RNase H derived by site-directed mutagenesis of a conserved histidine 539 are preferentially impaired in their exo- and less in their endonuclease activity. A specific polypurine-rich oligonucleotide created at the polypurine tract (PPT), serves as a primer for plus-strand DNA synthesis. Initiation of DNA synthesis at the PPT-primer can be demonstrated for the wt enzyme in vitro in contrast to the mt enzymes which do not recognize this sequence. A replication model based on these results is presented.
逆转录病毒复制酶逆转录酶（RT）与一种核糖核酸酶H相关联，该核糖核酸酶H能特异性水解RNA-DNA杂交体中的RNA。当用体外合成的病毒RNA与较短的合成DNA寡核苷酸杂交进行分析时，核糖核酸酶H表现出内切核酸酶和外切核酸酶活性。在脱氧核糖核苷酸存在的情况下，RT与核糖核酸酶H协同作用合成DNA，核糖核酸酶H同时水解RNA模板。通过对保守的组氨酸539进行定点诱变获得的核糖核酸酶H突变体，其外切核酸酶活性优先受损，内切核酸酶活性受损程度较小。在多聚嘌呤序列（PPT）处产生的一种特定的富含多聚嘌呤的寡核苷酸，可作为正链DNA合成的引物。与不识别该序列的突变酶相反，野生型酶在体外可证明在PPT引物处启动DNA合成。基于这些结果提出了一种复制模型。
