ROLE OF THE AVIAN RETROVIRAL PROTEASE IN THE ACTIVATION OF REVERSE-TRANSCRIPTASE DURING VIRION ASSEMBLY
ROLE OF THE AVIAN RETROVIRAL PROTEASE IN THE ACTIVATION OF REVERSE-TRANSCRIPTASE DURING VIRION ASSEMBLY
复制标题
DOI：
10.1128/jvi.65.11.6205-6217.1991
复制
发表时间：
1991-11-01
影响因子：
5.4
通讯作者：
WILLS, JW
中科院分区：
文献类型：
--
作者：
CRAVEN, RC;BENNETT, RP;WILLS, JW
禽肉瘤-白血病病毒群的逆转录病毒以两种前体形式合成它们的病毒蛋白酶（PR）——作为Gag前体的羧基末端结构域和作为Gag- pol前体的嵌入结构域。我们之前已经证明，Gag衍生的PR在没有嵌入PR的情况下完全能够处理Gag前体（R. P. Bennett， S. Rhee， R. C. Craven， E. Hunter， J. W. Wills， J. Virol. 65:272- 280,1991）。在这项研究中，我们研究了Gag-Pol的PR结构域是否在Pol蛋白的成熟中起重要作用的问题。Gag- pol前体在Gag缺失的情况下通过融合Gag和pol阅读框的基于猴病毒40的载体表达。融合蛋白在转染的细胞中积累到高水平，而不被释放到培养基中，但可以通过从第二个载体中共表达Gag蛋白而被拯救到颗粒中。所得到的颗粒含有成熟的Gag和Pol蛋白和活性逆转录酶（RT）。利用该互补系统，研究了Gag和/或Gag- pol蛋白中PR缺陷对RT活化的影响。结果表明，Gag前体上存在功能性PR，而Gag- pol上没有，这是完全激活RT所必需的。Gag- pol的嵌入PR不能对Gag前体进行任何可检测的处理，并且在没有功能性Gag PR结构域的情况下，只能将RT激活到低水平。最后，Gag-Pol PR结构域的一些点突变抑制了野生型PR在trans中对RT的激活，这表明Gag-Pol中PR结构域的正确构象是激活RT的先决条件。
The retroviruses of the avian sarcoma-leukosis virus group synthesize their viral protease (PR) in two precursor forms-as a carboxy-terminal domain of the Gag precursor and as an embedded domain within the Gag-Pol precursor. We have shown previously that the Gag-derived PR is fully capable of processing the Gag precursor in the absence of the embedded PR (R. P. Bennett, S. Rhee, R. C. Craven, E. Hunter, and J. W. Wills, J. Virol. 65:272-280, 1991). In this study, we examined the question of whether or not the PR domain of Gag-Pol has an essential role in the maturation of the Pol proteins. The Gag-Pol precursor was expressed in the absence of Gag by use of a simian virus 40-based vector in which the gag and pol reading frames were fused. The fusion protein accumulated to high levels in transfected cells without being released into the medium but could be rescued into particles by coexpression of the Gag protein from a second vector. The resulting particles contained mature Gag and Pol proteins and active reverse transcriptase (RT). Using this complementation system, the effects of PR defects in the Gag and/or Gag-Pol proteins on the activation of RT were examined. The results showed that the presence of a functional PR on the Gag precursor, but not on Gag-Pol, was required for full activation of RT. The embedded PR of Gag-Pol was unable to carry out any detectable processing of the Gag precursor and was able to activate RT to only a low level in the absence of a functional Gag PR domain. Finally, some point mutations in the Gag-Pol PR domain inhibited activation of RT in trans by a wild-type PR, suggesting that the correct conformation of the PR domain in Gag-Pol is prerequisite for activation of RT.