Kouno Takahide, Luengas Elizabeth M, Shigematsu Megumi, Shandilya Shivender M D, Zhang JingYing, Chen Luan, Hara Mayuko, Schiffer Celia A, Harris Reuben S, Matsuo Hiroshi
1] Biochemistry, Molecular Biology and Biophysics Department, Institute for Molecular Virology, University of Minnesota, Minneapolis, Minnesota, USA. [2] Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
Biochemistry, Molecular Biology and Biophysics Department, Masonic Cancer Center, Center for Genome Engineering, Institute for Molecular Virology, University of Minnesota, Minneapolis, Minnesota, USA.
Nat Struct Mol Biol. 2015 Jun;22(6):485-91. doi: 10.1038/nsmb.3033. Epub 2015 May 18.
The human APOBEC3G (A3G) DNA cytosine deaminase restricts and hypermutates DNA-based parasites including HIV-1. The viral infectivity factor (Vif) prevents restriction by triggering A3G degradation. Although the structure of the A3G catalytic domain is known, the structure of the N-terminal Vif-binding domain has proven more elusive. Here, we used evolution- and structure-guided mutagenesis to solubilize the Vif-binding domain of A3G, thus permitting structural determination by NMR spectroscopy. A smaller zinc-coordinating pocket and altered helical packing distinguish the structure from previous catalytic-domain structures and help to explain the reported inactivity of this domain. This soluble A3G N-terminal domain is bound by Vif; this enabled mutagenesis and biochemical experiments, which identified a unique Vif-interacting surface formed by the α1-β1, β2-α2 and β4-α4 loops. This structure sheds new light on the Vif-A3G interaction and provides critical information for future drug development.
人类载脂蛋白B mRNA编辑酶催化多肽样蛋白3G（APOBEC3G，A3G）DNA胞嘧啶脱氨酶可限制包括HIV-1在内的基于DNA的寄生虫并使其发生高度突变。病毒感染性因子（Vif）通过触发A3G降解来阻止这种限制作用。尽管A3G催化结构域的结构已为人所知，但事实证明，其N端Vif结合结构域的结构更难捉摸。在此，我们利用进化和结构导向诱变使A3G的Vif结合结构域溶解，从而能够通过核磁共振光谱法确定其结构。一个较小的锌配位口袋和改变的螺旋堆积将该结构与先前的催化结构域结构区分开来，并有助于解释该结构域已报道的无活性。这种可溶的A3G N端结构域与Vif结合；这使得诱变和生化实验成为可能，这些实验确定了由α1-β1、β2-α2和β4-α4环形成的独特的Vif相互作用表面。该结构为Vif-A3G相互作用提供了新的见解，并为未来的药物开发提供了关键信息。
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