Structural biology of host factors affecting retroviral integration

越来越清楚的是，参与逆转录病毒整合的重要宿主细胞蛋白是晶状体上皮衍生生长因子（LEDGF/p75），其是一种转录共激活因子，已显示其影响HIV整合酶的靶位点选择（Ciuffi et al.，2005;拉诺等人，2006年）。LEDGF直接与HIV整合酶相互作用，并已被提出作为连接前整合复合物与染色质的系链。最近，已经确定了LEDGF的整合酶结合区和HIV整合酶的催化结构域的共晶体结构（Cherepanov等人，2005年）。然而，目前还不清楚LEDGF如何与染色质相互作用并将整合酶引导至其作用位点。我们已经将我们的结构注意力集中在LEDGF的N-末端结构域上，LEDGF是PWWP蛋白家族的成员。已经假设该N-末端结构域与裸DNA结合，但最近的数据表明它可能与其他染色质元件结合（Botbol等人，2008年）。特别是，LEDGF的PWWP结构域和Tudor家族蛋白质之间的相似性向我们表明，它可能结合组蛋白尾部的甲基赖氨酸残基。我们已经表达和纯化了几个包含该结构域的构建体，并使用该结构域与从鸡红细胞纯化的核小体和修饰的肽进行结合研究。与DNA和肽复合的LEDGF PWWP结构域的结晶试验正在进行中。 Botbol，Y.，Raghavendra，N.K.，Rahman，S.，Engelman，A.，和Lavigne，M.等人（2008）Nucleic Acids Res.36，1237-1246。 Cherepanov，P.，安布罗西奥，A.L.B.，Rahman，S.，Ellenberger，T.，和Engelman，A.等人（2005）Proc. Acad. Sci. USA 102，17308-17313. Ciuffi，A.，等人（2005）Nature Med.11，1287-1289。 拉诺，M.，等人（2006）Science 314，461-464。

It has become increasingly clear that an important host cell protein involved in retroviral integration is lens-epithelium-derived growth factor (LEDGF/p75), a transcriptional coactivator that has been shown to influence target site selection by HIV integrase (Ciuffi et al., 2005; Llano et al., 2006). LEDGF interacts directly with HIV integrase, and has been proposed to act as a tether that links pre-integration complexes to chromatin. Recently, the co-crystal structure of the integrase-binding region of LEDGF and the catalytic domain of HIV integrase has been determined (Cherepanov et al., 2005). However, it is not yet understood how LEDGF interacts with chromatin and directs integrase to its site of action. We have focused our structural attention on the N-terminal domain of LEDGF, a member of the PWWP family of proteins. This N-terminal domain has been hypothesized to bind to naked DNA but recent data suggests that it may may bind to other chromatin elements (Botbol et al., 2008). In particular, the resemblance between LEDGF's PWWP domain and proteins in the Tudor family suggested to us that it may bind methyl lysine residues on histone tails. We have expressed and purified several constructs encompassing this domain, and have used this domain for binding studies with both nucleosomes purified from chicken erythrocytes and modified peptides. Crystallization trials of LEDGF PWWP domain complexed with DNA and peptides are underway. Botbol, Y., Raghavendra, N.K., Rahman, S., Engelman, A., and Lavigne, M. (2008) Nucleic Acids Res. 36, 1237-1246. Cherepanov, P., Ambrosio, A.L.B., Rahman, S., Ellenberger, T., and Engelman, A. (2005) Proc. Natl. Acad. Sci. USA 102, 17308-17313. Ciuffi, A., et al. (2005) Nature Med. 11, 1287-1289. Llano, M., et al. (2006) Science 314, 461-464.