van Gent D C, Vink C, Groeneger A A, Plasterk R H
Division of Molecular Biology, Netherlands Cancer Institute, Amsterdam.
EMBO J. 1993 Aug;12(8):3261-7. doi: 10.1002/j.1460-2075.1993.tb05995.x.
HIV integrase (IN) cleaves two nucleotides off the 3' end of viral DNA and integrates viral DNA into target DNA. Previously, three functional domains in the HIV IN protein have been identified: (i) the central catalytic domain, (ii) the C-terminal DNA binding domain, and (iii) the N-terminal region, which is also necessary for activity. We have now investigated whether IN proteins mutated in different domains can complement each other. Mutant D116I does not contain an intact active site, but does bind DNA, whereas the C-terminal deletion mutant C delta 73 does not bind DNA, but does have an intact active site. Neither mutant protein mediates site-specific cleavage or integration. However, a mixture of both proteins is active, suggesting that IN functions as an oligomer, and that two subunits can have different functions; one subunit binds the (viral) DNA and another subunit provides the active site. We found three classes of mutants, corresponding to the three domains mentioned above. Mutants from different classes, but not from the same class, can complement each other. However, complementation is most efficient when the N- and C-termini are present on the same molecule.
人类免疫缺陷病毒整合酶（IN）从病毒DNA的3'末端切除两个核苷酸，并将病毒DNA整合到靶DNA中。此前，已在HIV IN蛋白中鉴定出三个功能结构域：（i）中央催化结构域，（ii）C末端DNA结合结构域，以及（iii）N末端区域，其对活性也必不可少。我们现在研究了在不同结构域发生突变的IN蛋白是否能相互补充。突变体D116I不包含完整的活性位点，但能结合DNA，而C末端缺失突变体C delta 73不结合DNA，但具有完整的活性位点。两种突变蛋白均不介导位点特异性切割或整合。然而，两种蛋白的混合物具有活性，这表明IN作为一种寡聚体发挥作用，且两个亚基可具有不同功能；一个亚基结合（病毒）DNA，另一个亚基提供活性位点。我们发现了三类突变体，分别对应上述三个结构域。来自不同类别而非同一类别的突变体可以相互补充。然而，当N末端和C末端存在于同一分子上时，互补效率最高。
