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一个重要宿主细胞蛋白是透镜上皮衍生生长因子（LEDGF/p75），这是一种转录辅助激活因子，已被证明可以影响HIV整合酶的靶位点选择（Ciuffi等人，2005；Llano等人，2006）。LEDGF直接与HIV整合酶相互作用，并被认为是将整合前复合物与染色质连接起来的系绳。最近，已经确定了LEDGF整合酶结合区和HIV整合酶催化结构域的共晶结构（Cherepanov et al., 2005）。然而，目前尚不清楚LEDGF如何与染色质相互作用并将整合酶引导到其作用位点。我们将结构注意力集中在LEDGF的n端结构域上，LEDGF是PWWP蛋白家族的成员。这个n端结构域被假设与裸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 et al., 2008）。特别地，LEDGF的PWWP结构域与Tudor家族蛋白的相似性提示我们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包含该结构域的几个结构体，并将该结构域用于从鸡红细胞和修饰肽中纯化的核小体的结合研究。LEDGF PWWP结构域与DNA和肽络合的结晶试验正在进行中。

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.