THE EFFECTS OF DOMAIN SWAPPING IN HIV-1 CAPSID PROTEIN

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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.