Department of Pharmaceutical Sciences, College of Pharmacy, University of Nebraska Medical Center, Omaha, Nebraska, 68198-6025, USA.
Sci Rep. 2018 Dec 18;8(1):17953. doi: 10.1038/s41598-018-36372-6.
APOBEC3G (A3G) belongs to the family of cytosine deaminases that play an important role in the innate immune response. Similar to other, two-domain members of the APOBEC family, A3G is prone to concentration-dependent oligomerization, which is an integral for its function in the cell. It is shown that oligomerization of A3G is related to the packing mechanism into virus particle and, is critical for the so-called roadblock model during reverse transcription of proviral ssDNA. The role of oligomerization for deaminase activity of A3G is widely discussed in the literature; however, its relevance to deaminase activity for different oligomeric forms of A3G remains unclear. Here, using Atomic Force Microscopy, we directly visualized A3G-ssDNA complexes, determined their yield and stoichiometry and in parallel, using PCR assay, measured the deaminase activity of these complexes. Our data demonstrate a direct correlation between the total yield of A3G-ssDNA complexes and their total deaminase activity. Using these data, we calculated the relative deaminase activity for each individual oligomeric state of A3G in the complex. Our results show not only similar deaminase activity for monomer, dimer and tetramer of A3G in the complex, but indicate that larger oligomers of A3G retain their deaminase activity.
APOBEC3G（A3G）属于胞嘧啶脱氨酶家族，在先天免疫反应中发挥重要作用。与 APOBEC 家族的其他两个结构域成员相似，A3G 易于浓度依赖性寡聚化，这是其在细胞中功能的组成部分。研究表明，A3G 的寡聚化与病毒颗粒的包装机制有关，并且对前病毒 ssDNA 逆转录过程中的所谓阻碍模型至关重要。寡聚化对 A3G 脱氨酶活性的作用在文献中广泛讨论；然而，其对不同寡聚形式的 A3G 的脱氨酶活性的相关性尚不清楚。在这里，我们使用原子力显微镜直接可视化 A3G-ssDNA 复合物，确定了它们的产量和化学计量，并通过 PCR 测定平行测量这些复合物的脱氨酶活性。我们的数据表明 A3G-ssDNA 复合物的总产量与其总脱氨酶活性之间存在直接相关性。使用这些数据，我们计算了复合物中每个 A3G 寡聚体状态的相对脱氨酶活性。我们的结果表明，复合物中 A3G 的单体、二聚体和四聚体具有相似的脱氨酶活性，并且表明较大的 A3G 寡聚体保留其脱氨酶活性。
