Telesnitsky A, Goff S P
Department of Biochemistry and Molecular Biophysics, Columbia University College of Physicians and Surgeons, New York, NY 10032.
EMBO J. 1993 Nov;12(11):4433-8. doi: 10.1002/j.1460-2075.1993.tb06128.x.
Retroviral DNA synthesis requires both the DNA polymerase and the RNaseH activities of reverse transcriptase (RT). To test whether two defective RTs--one carrying a mutation in the RNaseH domain and the other with a mutation in DNA polymerase--could work together to complete viral DNA synthesis, we generated phenotypically mixed virions of Moloney murine leukemia virus (M-MuLV) that contained two kinds of mutant RTs. One RNaseH catalytic site mutant complemented both tested DNA polymerase mutants and small amounts of intact viral DNA were generated. This demonstrates that retroviral DNA synthesis can be completed--albeit inefficiently--when DNA polymerase and RNaseH activities are provided by separate RT molecules. Other RNaseH mutants failed to complement, suggesting that some aspects of the RNaseH domain are essential to RT's DNA polymerase function. Phenotypically mixed virions were also used to demonstrate that RT and integrase (IN) can be provided by separate polyprotein precursors and complete the early stages of retroviral replication.
逆转录病毒DNA合成需要逆转录酶（RT）的DNA聚合酶和核糖核酸酶H（RNaseH）活性。为了测试两个缺陷型RT（一个在RNaseH结构域携带突变，另一个在DNA聚合酶携带突变）是否能共同作用完成病毒DNA合成，我们构建了含有两种突变型RT的莫洛尼鼠白血病病毒（M-MuLV）表型混合病毒粒子。一种RNaseH催化位点突变体可互补两种测试的DNA聚合酶突变体，并产生了少量完整的病毒DNA。这表明，当DNA聚合酶和RNaseH活性由单独的RT分子提供时，逆转录病毒DNA合成可以完成，尽管效率不高。其他RNaseH突变体无法互补，这表明RNaseH结构域的某些方面对RT的DNA聚合酶功能至关重要。表型混合病毒粒子也被用于证明RT和整合酶（IN）可以由单独的多蛋白前体提供，并完成逆转录病毒复制的早期阶段。
