Activities and substrate specificity of the evolutionarily conserved central domain of retroviral integrase.
Activities and substrate specificity of the evolutionarily conserved central domain of retroviral integrase.
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逆转录病毒整合酶进化保守的中心域的活性和底物特异性。
DOI：
10.1016/s0042-6822(95)80060-3
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发表时间：
1995
期刊：
影响因子：
3.7
通讯作者：
Skalka,AM
中科院分区：
文献类型：
--
作者：
Kulkosky,J;Katz,RA;Merkel,G;Skalka,AM
逆转录病毒整合酶（inverse integrase，IN）是病毒编码的一种酶，它是病毒DNA插入宿主染色体所必需的，为了定位和确定这种独特的病毒酶的最小功能结构域的性质，构建了一系列Rous肉瘤病毒（Rous sarcoma virus，RSV）和人类免疫缺陷病毒（human immunodeficiency virus，HIV）IN的N端和C端缺失序列。首先测试RSV IN缺失突变体从代表病毒DNA末端的底物末端去除两个核苷酸的能力，以评估N和C区对该反应（称为加工）的贡献。结果表明，需要完整RSV蛋白的C-末端氨基酸来维持加工反应的特异性。虽然缺乏加工，RSV缺失突变体表现出二级内切核酸酶活性，这是无法区分的野生型IN，证明所有保留一些酶活性。RSV和一组较大的HIV-1、IN缺失突变体使用寡脱氧核苷酸底物进行分子内协同切割-连接反应的能力，该寡脱氧核苷酸底物模拟在共价闭合的最后步骤之前发现的中间病毒-宿主DNA连接。从这样的分析的复合结果定义了一个最小的功能中心区的140个氨基酸的每一种酶，包括高度保守的D，D（35）E结构域。HIV-1和HIV-2 IN的结果还表明，协同切割-连接的效率取决于反应位点处DNA底物的病毒组分内CA/GT碱基对的存在。甚至HIV-1 IN的分离的中心区域也表现出最佳活性的这种序列要求。我们的结论是，这种进化上保守的IN中心区域不仅编码的酶的催化活性所需的残基，但也窝藏一些或所有的决定因素负责识别的CA/GT二核苷酸，存在于所有逆转录病毒DNA的末端。
The retroviral integrase (IN) is a virus-encoded enzyme that is essential for insertion of viral DNA into the host chromosome.In order to map and define the properties of a minimal functional domain for this unique viral enzyme, a series of N- and C-terminal deletions of both Rous sarcoma virus (RSV) and human immunodeficiency virus (HIV) INs were constructed. The RSV IN deletion mutants were first tested for their ability to remove two nucleotides from the end of a substrate representing the terminus of viral DNA in order to assess the contribution of N and C regions towards this reaction, referred to as processing. The results suggest that C-terminal amino acids of the intact RSV protein are required to maintain specificity of the processing reaction. Though deficient for processing, the RSV deletion mutants exhibited a secondary endonucleolytic activity that was indistinguishable from that of wild-type IN, demonstrating that all retained some enzymatic activity. RSV, and a larger set of HIV-1, IN deletion mutants were then tested for their ability to perform an intramolecular, concerted cleavage- ligation reaction using an oligodeoxynucleotide substrate that mimics the intermediate viral-host DNA junction found prior to the final step of covalent closure. The composite results from such analyses define a minimal functional central region of ∼140 amino acids for each enzyme that includes the highly conserved D,D(35)E domain. Results with HIV-1 and HIV-2 IN also indicate that the efficiency of concerted cleavage- ligation depends upon the presence of CA/GT base pairs within the viral component of the DNA substrate at the reaction site. Even the isolated central region of HIV-1 IN exhibited this sequence requirement for optimal activity. We conclude that this evolutionarily conserved central region of IN not only encodes residues that are required for the catalytic activity of the enzyme but also harbors some or all of the determinants responsible for recognition of the CA/GT dinucleotides that are present at the ends of all retroviral DNAs.
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逆转录病毒整合酶以多聚体的形式发挥作用，并且可以催化翻转。
DOI：
--
发表时间：
1992
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Jones,KS;Coleman,J;Merkel,GW;Laue,TM;Skalka,AM
通讯作者：
Skalka,AM
不同域中突变的 HIV 整合酶蛋白之间的互补。
DOI：
10.1002/j.1460-2075.1993.tb05995.x
发表时间：
1993
期刊：
The EMBO Journal
影响因子：
--
作者：
D. V. Gent;Cornelis Vink;A. A. M. O. Groeneger;Ronald H. A. Plasterk
通讯作者：
Ronald H. A. Plasterk
影响因子：
3.7
作者：
DRELICH, M;WILHELM, R;MOUS, J
通讯作者：
MOUS, J
逆转录病毒 DNA 整合：转座子改组的经验教训。
影响因子：
3.5
作者：
Skalka,AM
通讯作者：
Skalka,AM
逆转录病毒整合酶的加工和连接活动需要保守的丝氨酸。
DOI：
10.1073/pnas.89.15.6741
发表时间：
1992
影响因子：
11.1
作者：
Katz,RA;Mack,JP;Merkel,G;Kulkosky,J;Ge,Z;Leis,J;Skalka,AM
通讯作者：
Skalka,AM