Lutzke R A, Vink C, Plasterk R H
Division of Molecular Biology, Netherlands Cancer Institute, Amsterdam.
Nucleic Acids Res. 1994 Oct 11;22(20):4125-31. doi: 10.1093/nar/22.20.4125.
The human immunodeficiency virus (HIV) integrase (IN) protein mediates an essential step in the retroviral lifecycle, the integration of viral DNA into human DNA. A DNA-binding domain of HIV IN has previously been identified in the C-terminal part of the protein. We tested truncated proteins of the C-terminal region of HIV-1 IN for DNA binding activity in two different assays: UV-crosslinking and southwestern blot analysis. We found that a polypeptide fragment of 50 amino acids (IN220-270) is sufficient for DNA binding. In contrast to full-length IN protein, this domain is soluble under low salt conditions. DNA binding of IN220-270 to both viral DNA and non-specific DNA occurs in an ion-independent fashion. Point mutations were introduced in 10 different amino acid residues of the DNA-binding domain of HIV-2 IN. Mutation of basic amino acid K264 results in strong reduction of DNA binding and of integrase activity.
人类免疫缺陷病毒（HIV）整合酶（IN）蛋白介导逆转录病毒生命周期中的一个关键步骤，即将病毒DNA整合到人类DNA中。HIV IN的一个DNA结合结构域先前已在该蛋白的C末端部分被鉴定出来。我们通过两种不同的检测方法测试了HIV-1 IN C末端区域的截短蛋白的DNA结合活性：紫外线交联和蛋白质印迹分析。我们发现一个50个氨基酸的多肽片段（IN220-270）足以进行DNA结合。与全长IN蛋白不同，该结构域在低盐条件下是可溶的。IN220-270与病毒DNA和非特异性DNA的DNA结合以离子独立的方式发生。在HIV-2 IN的DNA结合结构域的10个不同氨基酸残基中引入了点突变。碱性氨基酸K264的突变导致DNA结合和整合酶活性的强烈降低。
