THE EFFECTS OF DOMAIN SWAPPING IN HIV-1 CAPSID PROTEIN

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 HIV-1的组装是一个多步骤的过程，在这个过程中，几个单独的结构蛋白经历了实质性的形态重排。衣壳蛋白(CA)在这个组装过程中起着至关重要的作用，但人们对此知之甚少。CA含有两个独立折叠的结构域，N-末端(NTD)和C-末端(CTD)结构域，由灵活的连接子连接。HIV-1的CTD已经被证明在溶液中在成熟的衣壳蛋白和分离的结构域中都是二聚的(Gamble等人。1997年)。目前成熟病毒粒子的结构模型表明，成熟核心的基本构件是由NTD通过CTD的二聚化而自结合形成的CA六角体(Gamble等人)。1997年)。此外，已经证明这种六聚体的形成需要CTD二聚化(Lanman等人)。2003年)。 HIV的CTD包含主要同源区(MHR)，这是一个由20个氨基酸组成的序列，在不同的逆转录病毒属中高度保守(Wills和Craven 1991)。众所周知，病毒组装对MHR突变高度敏感，但从结构角度看，MHR突变如此有害的原因尚不清楚(凯恩斯和克雷文2001)。最近的研究表明，CA的CTD和二聚体锌指相关结构域扫描之间存在结构同源性(Ivanov等人)。2005)。然而，与CA的CTD相比，扫描结构域二聚体被结构域交换。在结构域交换的二聚体中，来自一个亚基的结构元件与来自另一个亚基的相应结构元件交换(Liu和Eisenberg 2002)。扫描二聚体的结构域交换区域对应于CTD的MHR。结构域交换的二聚体将为MHR的重要性提供解释，因为这一区域的氢键将通过二聚体界面发生。为了确定当C-末端结构域被交换时CA是否可以组装，设计了一个融合结构，其中HIV-1 CA的C-末端结构域被扫描结构域取代。 这些相互作用将用H/D交换和快速光化学氧化来研究。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The assembly of HIV-1 is a multi-step process in which several individual structural proteins undergo a substantial morphological rearrangement. The capsid (CA) protein plays a crucial but poorly understood role in this assembly. CA contains two-independently folded domains, the N-terminal (NTD) and C-terminal (CTD) domains, connected by a flexible linker. The CTD of HIV-1 has been shown to dimerize in solution both in the mature capsid protein and in the isolated domain (Gamble et al. 1997). The current structural model of the mature virion indicates the fundamental building block of the mature core is attributed to a CA hexamer formed by the self-association of the NTD tied together by dimerization of the CTD (Gamble et al. 1997). Additionally, CTD dimerization has been shown to be required for this hexamer formation (Lanman et al. 2003). The CTD of HIV contains the major homology region (MHR), a sequence of 20 amino acids that is highly conserved across different genera of retroviruses (Wills and Craven 1991). It is well documented that viral assembly is highly sensitive to MHR mutations but from a structural perspective it is not clear why MHR mutations are so deleterious (Cairns and Craven 2001). Recent studies have shown that a structural homology exists between the CTD of CA and the dimeric zinc finger associated domain SCAN (Ivanov et al. 2005). However, the SCAN domain dimer is domain swapped in comparison to the CTD of CA. In a domain swapped dimer, a structural element from one subunit is exchanged with the corresponding structural element from another subunit (Liu and Eisenberg 2002). The domain swapped region of the SCAN dimer corresponds to the MHR of the CTD. A domain swapped dimer would provide an explanation for the importance of the MHR because hydrogen bonding in this region would occur across the dimer interface. To determine whether CA can assemble when the C-terminal domain is domain swapped a fusion construct was engineered where the C-terminal domain of HIV-1 CA was replaced with the SCAN domain. These interactions will be studied using H/D exchange and fast photochemical oxidation.