Summers M F, Henderson L E, Chance M R, Bess J W, South T L, Blake P R, Sagi I, Perez-Alvarado G, Sowder R C, Hare D R
Department of Chemistry and Biochemistry, University of Maryland Baltimore County 21228.
Protein Sci. 1992 May;1(5):563-74. doi: 10.1002/pro.5560010502.
All retroviral nucleocapsid (NC) proteins contain one or two copies of an invariant 3Cys-1His array (CCHC = C-X2-C-X4-H-X4-C; C = Cys, H = His, X = variable amino acid) that are essential for RNA genome packaging and infectivity and have been proposed to function as zinc-binding domains. Although the arrays are capable of binding zinc in vitro, the physiological relevance of zinc coordination has not been firmly established. We have obtained zinc-edge extended X-ray absorption fine structure (EXAFS) spectra for intact retroviruses in order to determine if virus-bound zinc, which is present in quantities nearly stoichiometric with the CCHC arrays (Bess, J.W., Jr., Powell, P.J., Issaq, H.J., Schumack, L.J., Grimes, M.K., Henderson, L.E., & Arthur, L.O., 1992, J. Virol. 66, 840-847), exists in a unique coordination environment. The viral EXAFS spectra obtained are remarkably similar to the spectrum of a model CCHC zinc finger peptide with known 3Cys-1His zinc coordination structure. This finding, combined with other biochemical results, indicates that the majority of the viral zinc is coordinated to the NC CCHC arrays in mature retroviruses. Based on these findings, we have extended our NMR studies of the HIV-1 NC protein and have determined its three-dimensional solution-state structure. The CCHC arrays of HIV-1 NC exist as independently folded, noninteracting domains on a flexible polypeptide chain, with conservatively substituted aromatic residues forming hydrophobic patches on the zinc finger surfaces. These residues are essential for RNA genome recognition, and fluorescence measurements indicate that at least one residue (Trp37) participates directly in binding to nucleic acids in vitro. The NC is only the third HIV-1 protein to be structurally characterized, and the combined EXAFS, structural, and nucleic acid-binding results provide a basis for the rational design of new NC-targeted antiviral agents and vaccines for the control of AIDS.
所有逆转录病毒核衣壳（NC）蛋白都包含一到两个不变的3个半胱氨酸 - 1个组氨酸序列（CCHC = C-X2-C-X4-H-X4-C；C = 半胱氨酸，H = 组氨酸，X = 可变氨基酸），这些序列对于RNA基因组包装和感染性至关重要，并且被认为起着锌结合结构域的作用。尽管这些序列在体外能够结合锌，但锌配位的生理相关性尚未得到确凿证实。我们已获得完整逆转录病毒的锌边缘扩展X射线吸收精细结构（EXAFS）光谱，以确定与病毒结合的锌（其含量与CCHC序列几乎化学计量存在，Bess, J.W., Jr., Powell, P.J., Issaq, H.J., Schumack, L.J., Grimes, M.K., Henderson, L.E., & Arthur, L.O., 1992, J. Virol. 66, 840 - 847）是否存在于独特的配位环境中。所获得的病毒EXAFS光谱与具有已知3个半胱氨酸 - 1个组氨酸锌配位结构的模型CCHC锌指肽的光谱非常相似。这一发现与其他生化结果相结合，表明在成熟逆转录病毒中，大部分病毒锌与NC CCHC序列配位。基于这些发现，我们扩展了对HIV - 1 NC蛋白的核磁共振研究，并确定了其三维溶液态结构。HIV - 1 NC的CCHC序列以独立折叠、不相互作用的结构域形式存在于柔性多肽链上，保守取代的芳香族残基在锌指表面形成疏水斑块。这些残基对于RNA基因组识别至关重要，荧光测量表明至少一个残基（Trp37）在体外直接参与与核酸的结合。NC是第三个进行结构表征的HIV - 1蛋白，综合EXAFS、结构和核酸结合结果为合理设计用于控制艾滋病的新型靶向NC的抗病毒药物和疫苗提供了基础。
