APOBEC3 STRUCTURAL STUDIES

 描述（由申请人提供）：抗病毒APOBEC 3酶的发现被认为是HIV/AIDS分子病毒学中最有治疗前景的突破之一。几种APOBEC 3有可能通过整合到组装的病毒颗粒中，物理干扰逆转录的进程，以及将病毒cDNA胞嘧啶脱氨基为尿嘧啶来限制HIV复制。后一种抗病毒活性是APOBEC 3介导的限制性酶切的定义标志，解释了在患者来源的病毒序列中经常观察到的基因组链G至A突变。然而，这些有效的抗病毒活性被HIV病毒体感染因子（Vif）抵消，其与CBF-β异二聚化以形成降解APOBEC 3酶的E3泛素连接酶复合物。在这里，我们将解决这一领域的两个长期存在的问题。首先，我们将确定APOBEC 3结合的Vif/CBF-beta泛素连接酶复合物的X射线结构。这些结构将提供深入了解APOBEC 3-Vif结合机制，构象变化和这些宿主-病原体复合物的整体组织。其次，我们将阐明APOBEC 3/单链DNA复合物的X射线结构。这些结构将提供深入了解酶催化，DNA底物识别，DNA结合和催化活性释放之间的耦合机制。我们预计，APOBEC 3/Vif/CBF-beta和APOBEC 3/单链DNA大分子复合物的结构信息将在长期内非常重要，因为该领域将继续朝着药物化这些相互作用并通过天然先天免疫实现有效的HIV-1限制。

 DESCRIPTION (provided by applicant): The discovery of the antiviral APOBEC3 enzymes is regarded as one of the most therapeutically promising breakthroughs in HIV/AIDS molecular virology. Several APOBEC3s have the potential to restrict HIV replication by incorporating into assembling viral particles, physically interfering with the progression of reverse transcription, and deaminating viral cDNA cytosines to uracils. The latter antiviral activity is the defining hallmark of APOBEC3-mediated restriction, explaining the genomic strand G-to-A mutations that are frequently observed in patient-derived viral sequences. However, these potent antiviral activities are counteracted by the HIV virion infectivity factor (Vif), which heterodimerizes with CBF-beta in order to form an E3 ubiquitin ligase complex that degrades APOBEC3 enzymes. Here, we will address two persisting problems in this field. First, we will determine x-ray structures of APOBEC3-bound Vif/CBF-beta ubiquitin ligase complexes. These structures will provide insights into the APOBEC3-Vif binding mechanism, conformational changes, and the overall organization of these host-pathogen complexes. Second, we will elucidate X-ray structures of APOBEC3/single-stranded DNA complexes. These structures will provide insights into the mechanisms of enzymatic catalysis, DNA substrate recognition, and coupling between DNA-binding and the release of catalytic activity. We anticipate that structural information from both APOBEC3/Vif/CBF-beta and APOBEC3/single-stranded DNA macromolecular complexes will be important in the longer-term as the field continues to move toward drugging these interactions and enabling potent HIV-1 restriction through natural innate immunity.