Ordered processing of the human immunodeficiency virus type 1 GagPol precursor is influenced by the context of the embedded viral protease
Ordered processing of the human immunodeficiency virus type 1 GagPol precursor is influenced by the context of the embedded viral protease
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DOI：
10.1128/jvi.79.16.10601-10607.2005
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发表时间：
2005-08-01
影响因子：
5.4
通讯作者：
Kaplan, AH
中科院分区：
文献类型：
--
作者：
Pettit, SC;Clemente, JC;Kaplan, AH
人类免疫缺陷病毒I型（HIV-1） GagPol多蛋白前体被病毒编码的蛋白酶有序和准确地加工是感染性病毒颗粒适当组装不可或缺的步骤。HIV-1蛋白酶（PR）是一种99个氨基酸的酶，作为GagPol前体的一部分被翻译。之前，我们已经证明了前体加工中的初始事件是由cis中GagPol中的PR结构域完成的，然后才从多蛋白中释放出来。尽管前体的有序加工在病毒复制中起着关键作用，但定义切割顺序的力量仍然知之甚少。利用嵌入的PR处理全长HIV-1 GagPol的体外实验，我们检测了PR环境（嵌入在GagPol中与成熟的99氨基酸酶相比）对前体处理的影响。我们的数据表明，GagPol中的PR结构域切割前体中一些加工位点的能力受到限制。此外，我们发现这种限制取决于脯氨酸作为嵌入PR中的初始氨基酸的存在；当前体被嵌入的而不是成熟的PR加工时，在这个位置替换丙氨酸会在其他位点上产生增强的切割。总的来说，我们的数据支持一个模型，其中加工位点的选择和前体加工的顺序至少部分由GagPol本身的结构定义。
Ordered and accurate processing of the human immunodeficiency virus type I (HIV-1) GagPol polyprotein precursor by a virally encoded protease is an indispensable step in the appropriate assembly of infectious viral particles. The HIV-1 protease (PR) is a 99-amino-acid enzyme that is translated as part of the GagPol precursor. Previously, we have demonstrated that the initial events in precursor processing are accomplished by the PR domain within GagPol in cis, before it is released from the polyprotein. Despite the critical role that ordered processing of the precursor plays in viral replication, the forces that define the order of cleavage remain poorly understood. Using an in vitro assay in which the full-length HIV-1 GagPol is processed by the embedded PR, we examined the effect of PR context (embedded within GagPol versus the mature 99-amino-acid enzyme) on precursor processing. Our data demonstrate that the PR domain within GagPol is constrained in its ability to cleave some of the processing sites in the precursor. Further, we find that this constraint is dependent upon the presence of a proline as the initial amino acid in the embedded PR; substitution of an alanine at this position produces enhanced cleavage at additional sites when the precursor is processed by the embedded, but not the mature, PR. Overall, our data support a model in which the selection of processing sites and the order of precursor processing are defined, at least in part, by the structure of GagPol itself.