Nano-lmaging APOBECS Interactions

APOBECSG(A3G)和APOBECSF(A3F)参与了各种抗病毒蛋白质和核酸交易。重要的主要相互作用包括蛋白质间相互作用、结合单链核酸(RNA和单链DNA)以及HIV Vif的中和作用。充分阐明这些分子间的相互作用对于了解这些细胞蛋白抑制HIV复制的机制是必要的。齐聚在其活性中的作用是一个非常重要的问题。这些都是纳米级的相互作用，原子力显微镜是能够解决上述问题的选择方法。原子力显微镜目前的发展水平使这一独特的仪器能够提供丰富的重要信息，以表征纳米级的生物分子复合体。分子体系的形貌分析是原子力显微镜应用的初始领域，用于表征具有广泛复杂性的蛋白质-DNA体系的结构和特异性。除了成像，AFM还能够直接测量系统内的分子相互作用，这十年的研究导致了AFM在这一领域的应用取得了戏剧性的进展。我们的初步研究已经使我们能够提供A3G与单链DNA底物结合的第一张图像。我们还预计，所有形式的AFM技术的使用将帮助我们详细说明这些复合体，剖析蛋白质和化学要求，并将关键观察扩展到ASF。为了帮助实现这些首要目标，我们将实现以下三个具体目标：目标1.使用原子力显微镜以纳米分辨率成像A3复合体(计划目标1和2)；目标2.使用力光谱测量A3-ssDNA复合体内的相互作用(计划目标1和2)和目标3.使用AFM成像和探测模式表征A3-VIF相互作用(计划目标3)。该项目下的研究是我们总体计划项目的组成部分，旨在提供对A3-AS3、A3-核酸和A3-Vif相互作用的全面了解。 纳米规模的研究将提供一个平台，通过利用AS-VIF相互作用来评估任何未来发挥作用的疗法。

APOBECSG (A3G) and APOBECSF (A3F) are involved in various anti-viral protein and nucleic acid transactions. Important primary interactions include interprotein interactions, binding single-stranded nucleic acids (RNA and ssDNA), and neutralization by HIV Vif. The full elucidation of these intermolecular interactions is necessary to understand the mechanism by which these cellular proteins inhibit HIV replication. The role of oligomerization in their activity is an issue of great importance. These are interactions at the nanoscale level, and AFM is the method of choice capable to addressing problems listed above. AFM currently reached a level of development that enables this unique instrument to provide a wealth of important information to the characterization of biomolecular complexes at nanoscale. The topographic analysis of molecular system is the initial area of the AFM application for characterization of the structure and specificity of protein-DNA systems of a broad complexity. In addition to imaging, AFM is capable of direct measurements of molecular interaction within the system and research during this decade led to a dramatic progress in this area of the AFM applications. Our Preliminary Studies have already enabled us to provide the first images of A3G bound to ssDNA substrates. We further anticipate that the use of all modalities of AFM technology will help us detail these complexes, dissect the protein and chemical requirements, and extend key observations to ASF. To help achieve these overarching objectives, we will pursue the following three Specific Aims: Aim 1. Image A3 complexes at nanoscale resolution using AFM (Program Objectives 1 & 2); Aim 2. Measure interactions within A3-ssDNA complexes using force spectroscopy (Program Objectives 1 & 2) and Aim 3. Characterize A3-Vif interactions using AFM imaging and probing modalities (Program Objectives 3). The studies under this project are an integral component of our overall Program Project aiming to provide a comprehensive understanding of A3-AS3 A3-nucleic acid, and A3-Vif interactions. The nanoscale studies will provide a platform for evaluating any future therapeutics that function by leveraging the AS-Vif interaction.