整合病毒
中文名称
通路描述
逆转录病毒的 DNA 必须共价整合到宿主细胞染色体中才能指导子代病毒颗粒的产生（参见 Coffin 等人 1997 年；Hansen 等人 1998 年）。对突变体的分析确定了逆转录病毒 pol 基因中的整合酶编码区是整合过程所必需的（Donehower 1988；Donehower 和 Varmus 1984；Panganiban 和 Temin 1984；Quinn 和 Grandgenett 1988；Schwartzberg 等人 1984）。此外，逆转录病毒长末端重复序列（LTR）末端也是整合酶蛋白的识别位点，对于整合过程至关重要（Colicelli 和 Goff 1985, 1988；Panganiban 和 Temin 1983）。病毒基因组 RNA 被逆转录形成线性双链 DNA 分子，这是整合前病毒的前体（Brown 等人 1987, 1989；Fujiwara 和 Mizuuchi 1988）。前病毒与未整合的线性病毒 DNA 平行（Dhar 等人 1980；Hughes 等人 1978），但与逆转录产物不同，它缺少两端各两个碱基（Hughes 等人 1981）。整合 HIV 前病毒两侧是直接重复的细胞 DNA，长度为 5 个碱基对（Vincent 等人 1990）。这种在病毒 DNA 两侧重复的细胞序列的复制是整合机制的后果（Coffin 等人，1997）。线性病毒 DNA 在感染细胞的细胞质中与蛋白质形成复合物。这些复合物（称为“前整合复合物”，PICs）可以分离并证明它们在体外介导病毒 DNA 整合到靶 DNA 中（Bowerman 等人 1989；Brown 等人 1987；Ellison 等人 1990；Farnet 和 Haseltine 1990, 1991）。纯化的整合酶体外 assay 的开发允许阐明其酶学功能。前病毒由整合酶催化的两个反应形成：末端切割和链转移。使用纯化的整合酶的研究表明，它足以催化 3' 末端切割（Bushman 和 Craigie 1991；Craigie 等人 1990；Katzman 等人 1989；Sherman 和 Fyfe 1990）以及病毒 DNA 与细胞染色体或裸露靶 DNA 的连接（Bushman 等人 1990；Craigie 等人 1990；Katz 等人 1990）。HIV 整合酶催化从每个病毒 DNA 链的 3' 末端移除两个碱基，留下凹陷的 3'羟基基团（Brown 等人 1989；Fujiwara 和 Mizuuchi 1988；Roth 等人 1989；Sherman 和 Fyfe 1990）。这种末端切割反应对于正确的整合是必需的。它可能允许病毒从病毒 DNA 末端创建标准末端，这些末端可能由于逆转录酶的末端转移酶活性而具有异质性（Miller 等人 1997；Patel 和 Preston 1994）。此外，末端切割步骤与整合酶 -DNA 复合物的形成有关联（Ellison 和 Brown 1994；Vink 等人 1994）。末端切割后，暴露出一个羟基，该羟基位于 CA 二核苷酸之后。内部 LTR 位点对于整合也很重要（Balakrishnan 和 Jonsson 1997；Bushman 和 Craigie 1990；Leavitt 等人 1992）。在末端处理后，整合酶催化病毒 DNA 末端的羟基与宿主细胞 DNA 突出的 5'磷酸基团共价连接（Brown 等人 1987；Brown 等人 1989；Fujiwara 和 Mizuuchi 1988）。参与整合的 DNA 切割和连接反应如图示。病毒 DNA 3'末端切割和链转移反应均由单步转酯化化学介导，如反应产物的立体化学分析所示（Engelman 等人 1991）。纯化整合酶的生化分析表明，它需要二价金属离子——要么是 Mg2+，要么是 Mn2+——来与模型底物进行反应，这很可能介导反应化学（Bushman 和 Craigie 1991；Craigie 等人 1990；Katzman 等人 1989；Sherman 和 Fyfe 1990；Gao 等人 2004）。
英文描述
Integration of provirus For retroviral DNA to direct production of progeny virions it must become covalently integrated into the host cell chromosome (reviewed in Coffin et al. 1997; Hansen et al. 1998). Analyses of mutants have identified the viral integrase coding region (part of the retroviral pol gene) as essential for the integration process (Donehower 1988; Donehower and Varmus 1984; Panganiban and Temin 1984; Quinn and Grandgenett 1988; Schwartzberg et al. 1984). Also essential are regions at the ends of the viral long terminal repeats (LTRs) that serve as recognition sites for integrase protein (Colicelli and Goff 1985, 1988; Panganiban and Temin 1983).The viral genomic RNA is reverse transcribed to form a linear double-stranded DNA molecule, the precursor to the integrated provirus (Brown et al. 1987, 1989; Fujiwara and Mizuuchi 1988). The provirus is colinear with unintegrated linear viral DNA (Dhar et al. 1980; Hughes et al. 1978) but differs from the reverse transcription product in that it is missing two bases from each end (Hughes et al. 1981). Flanking the integrated HIV provirus are direct repeats of the cellular DNA that are 5 base pairs in length (Vincent et al. 1990). This duplication of cellular sequences flanking the viral DNA is generated as a consequence of the integration mechanism (Coffin et al., 1997). Linear viral DNA is found in a complex with proteins in the cytoplasm of infected cells. These complexes (termed "preintegration complexes", PICs) can be isolated and have been shown to mediate integration of viral DNA into target DNA in vitro (Bowerman et al. 1989; Brown et al. 1987; Ellison et al. 1990; Farnet and Haseltine 1990, 1991). The development of in vitro assays with purified integrase has allowed its enzymatic functions to be elucidated. The provirus is formed by two reactions catalyzed by the viral integrase: terminal cleavage and strand transfer. Studies with purified integrase have shown that it is sufficient for both 3' end cleavage (Bushman and Craigie 1991; Craigie et al. 1990; Katzman et al. 1989; Sherman and Fyfe 1990) and joining of the viral DNA to the cellular chromosome or naked target DNA (Bushman et al. 1990; Craigie et al. 1990; Katz et al. 1990). HIV integrase catalyze the removal of two bases from the 3' end of each viral DNA strand, leaving recessed 3' hydroxyl groups (Brown et al. 1989; Fujiwara and Mizuuchi 1988; Roth et al. 1989; Sherman and Fyfe 1990). This terminal cleavage reaction is required for proper integration. It may allow the virus to create a standard end from viral DNA termini that can be heterogeneous due to the terminal transferase activity of reverse transcriptase (Miller et al. 1997; Patel and Preston 1994). In addition, the terminal cleavage step is coupled to the formation of a stable integrase-DNA complex (Ellison and Brown 1994; Vink et al. 1994). Following terminal cleavage, a recessed hydroxyl is exposed that immediately follows a CA dinucleotide. More internal LTR sites are also important for integration (Balakrishnan and Jonsson 1997; Bushman and Craigie 1990; Leavitt et al. 1992). After end processing, integrase catalyzes the covalent attachment of hydroxyl groups at the viral DNA termini to protruding 5' phosphoryl ends of the host cell DNA (Brown et al. 1987; Brown et al. 1989; Fujiwara and Mizuuchi 1988). The DNA cleavage and joining reactions involved in integration are shown in the figure below. Both the viral DNA 3' end cleavage and strand transfer reactions are mediated by single-step transesterification chemistry as shown by stereochemical analysis of reaction products (Engelman et al. 1991). Biochemical analysis of purified integrase revealed that it requires a divalent metal - either Mg2+ or Mn2+ - to carry out reactions with model substrates, that probably mediate the reaction chemistry (Bushman and Craigie 1991; Craigie et al. 1990; Katzman et al. 1989; Sherman and Fyfe 1990; Gao et al. 2004).
所含基因
6 个基因