A soluble active mutant of HIV-1 integrase - Involvement of both the core and carboxyl-terminal domains in multimerization
A soluble active mutant of HIV-1 integrase - Involvement of both the core and carboxyl-terminal domains in multimerization
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DOI：
10.1074/jbc.271.13.7712
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发表时间：
1996-03-29
影响因子：
4.8
通讯作者：
Craigie, R
中科院分区：
文献类型：
--
作者：
Jenkins, TM;Engelman, A;Craigie, R
人类免疫缺陷病毒1型（HIV-1）整合酶的结构研究一直受到蛋白质溶解度低的阻碍。通过疏水残基的系统替换，我们先前确定了一个单一的氨基酸变化（F185 K），显着提高了HIV-1整合酶的催化结构域的溶解度，并使结构能够通过X射线晶体学确定。我们已经将相同的突变引入全长HIV-1整合酶。产生的重组蛋白在体外是可溶的且完全有活性，而携带突变的HIV-1由于病毒组装不当而具有复制缺陷。通过凝胶过滤和沉降平衡对重组蛋白的分析表明二聚体-四聚体自缔合。我们发现，参与多聚化的区域映射到催化核心和羧基末端结构域。这种蛋白质的溶解性显著提高，使其成为结构研究的良好候选者。
Structural studies of human immunodeficiency virus type 1 (HIV-1) integrase have been impeded by the low solubility of the protein. By systematic replacement of hydrophobic residues, we previously identified a single amino acid change (F185K) that dramatically improved the solubility of the catalytic domain of HIV-1 integrase and enabled the structure to be determined by x-ray crystallography. We have introduced the same mutation into full-length HIV-1 integrase. The resulting recombinant protein is soluble and fully active in vitro, whereas, HIV-1 carrying the mutation is replication-defective due to improper virus assembly. Analysis of the recombinant protein by gel filtration and sedimentation equilibrium demonstrate a dimer-tetramer self-association. We find that the regions involved in multimerization map to both the catalytic core and carboxyl-terminal domains. The dramatically improved solubility of this protein make it a good candidate for structural studies.