Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, Ottawa, Ontario, Canada.
Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, Ottawa, Ontario, Canada.
Virology. 2015 Sep;483:141-8. doi: 10.1016/j.virol.2015.04.019. Epub 2015 May 15.
APOBEC3G (A3G) is a host-expressed protein that inactivates retroviruses through the mutagenic deamination of cytosines (C) to uracils (U) in single-stranded DNA (ssDNA) replication products. A3G prefers to deaminate cytosines preceded by a cytosine (5'CC), whereas all other A3 proteins target cytosines in a 5'TC motifs. Structural and mutational studies have mapped the dinucleotide deamination preference of A3G to residues in loop 7 of the catalytic C-terminal domain of the protein. Here we report that A3G with a double W94A/W127A substitution in its N-terminus, designed to abolish RNA binding and protein oligomerization, alters the DNA deamination specificity of the enzyme from 5'CC to 5'TC on proviral DNA. We also show that the double substitution severely impairs its deaminase and antiretroviral activities on Vif-deficient HIV-1. Our results highlight that the N-terminal domain of the full length A3G protein has an important influence on its DNA sequence specificity and mutator activity.
APOBEC3G (A3G) 是一种宿主表达的蛋白，通过单链 DNA (ssDNA) 复制产物中胞嘧啶 (C) 的致突变脱氨作用转化为尿嘧啶 (U)，从而使逆转录病毒失活。A3G 更喜欢脱氨作用于紧随其后的胞嘧啶 (5'CC)，而其他所有 A3 蛋白则靶向 5'TC 基序中的胞嘧啶。结构和突变研究将 A3G 的二核苷酸脱氨作用偏好性映射到蛋白催化 C 端结构域环 7 中的残基上。在这里，我们报告说，A3G 的 N 端双 W94A/W127A 取代设计用于破坏 RNA 结合和蛋白寡聚化，使酶在原病毒 DNA 上的 DNA 脱氨特异性从 5'CC 改变为 5'TC。我们还表明，这种双重取代严重损害了它在缺乏 Vif 的 HIV-1 上的脱氨酶和抗逆转录病毒活性。我们的结果强调了全长 A3G 蛋白的 N 端结构域对其 DNA 序列特异性和突变活性有重要影响。
