Structural biology of host factors affecting retroviral integration

影响逆转录病毒整合的宿主因素的结构生物学

• 批准号: 8148764
• 负责人: Frederick Dyda
• 金额: $ 14.57万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8148764
• 关键词: Acquired Immunodeficiency Syndrome; Affect; Binding; Crystallization; Cytoplasm; DNA; Data; Elements; Erythrocytes; Family; HIV Integrase; Integration Host Factors; PWWP Domain; Protein Family; Transcription Coactivator; structural biology

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.

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），它是一种转录共激活因子，已被证明可以影响 HIV 整合酶的靶位点选择（Ciuffi 等人，2005 年；Llano 等人，2006 年）。 LEDGF 直接与 HIV 整合酶相互作用，并被提议充当连接预整合复合物与染色质的系绳。最近，LEDGF整合酶结合区和HIV整合酶催化结构域的共晶结构已被确定(Cherepanov等，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至其作用位点。我们将结构注意力集中在 LEDGF（PWWP 蛋白质家族成员）的 N 端结构域上。假设该 N 末端结构域与裸露 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）。特别是，LEDGF 的 PWWP 结构域与 Tudor 家族蛋白质之间的相似性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包含该结构域的几种构建体，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。 LEDGF PWWP 结构域与 DNA 和肽复合的结晶试验正在进行中。 Botbol, Y.、Raghavendra, N.K.、Rahman, S.、Engelman, A. 和 Lavigne, M. (2008) 核酸研究。 36、1237-1246。 Cherepanov, P.、Ambrosio, A.L.B.、Rahman, S.、Ellenberger, T. 和 Engelman, A. (2005) Proc。国家。阿卡德。科学。美国 102，17308-17313。 丘菲，A.，等人。 （2005）自然医学。 11、1287-1289。 拉诺，M.，等人。 (2006) 科学 314, 461-464。