Programa de Virología, Instituto de Ciencias Biomédicas, Facultad de Medicina, Universidad de Chile, Independencia 1027, Santiago, Chile.
Arch Biochem Biophys. 2010 Mar 1;495(1):28-34. doi: 10.1016/j.abb.2009.12.018. Epub 2009 Dec 21.
X-ray diffraction data on a few retroviral integrases show a flexible loop near the active site. By sequence alignment, the peptide region 207-218 of Mo-MLV IN appears to correspond to this flexible loop. In this study, residues H208, Y211, R212, Q214, S215 and S216 of Mo-MLV IN were mutated to determine their role on enzyme activity. We found that Y211A, R212A, R212K and Q214A decreased integration activity, while disintegration and 3'-processing were not significantly affected. By contrast H208A was completely inactive in all the assays. The core domain of Mo-MLV integrase was modeled and the flexibility of the region 207-216 was analyzed. Substitutions with low integration activity showed a lower flexibility than wild type integrase. We propose that the peptide region 207-216 is a flexible loop and that H208, Y211, R212 and Q214 of this loop are involved in the correct assembly of the DNA-integrase complex during integration.
X 射线衍射数据显示少数逆转录病毒整合酶在活性位点附近存在一个柔性环。通过序列比对，Mo-MLV IN 的肽段区域 207-218 似乎对应于这个柔性环。在这项研究中，我们对 Mo-MLV IN 的残基 H208、Y211、R212、Q214、S215 和 S216 进行了突变，以确定它们对酶活性的作用。我们发现 Y211A、R212A、R212K 和 Q214A 降低了整合活性，而解体和 3'-加工没有受到显著影响。相比之下，H208A 在所有测定中完全没有活性。我们对 Mo-MLV 整合酶的核心结构域进行了建模，并分析了该区域 207-216 的灵活性。整合活性较低的突变体显示出比野生型整合酶更低的灵活性。我们提出肽段区域 207-216 是一个柔性环，该环中的 H208、Y211、R212 和 Q214 参与了整合过程中 DNA-整合酶复合物的正确组装。
