DOMAIN MAPPING HIV VIF COMPLEXES BY LIMITED PROTEOLYSIS AND MASS-SPECTROMETRY

University Of California, San Francisco, San Francisco CA

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Abstract

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. The APOBEC3 family of enzymes protects primates from retroviruses such as HIV by hyperediting the viral genome thereby restricting viral spread in the host. HIV Vif combats this defense by binding APOBEC3 family members and hijacking a cellular complex that catalyzes the addition of the small protein Ubiquitin to APOBEC3, sentencing the APOBEC3 proteins for destruction and allowing viral spread. We have recently reconstituted the VIF Ubiquitin ligase from recombinant proteins and seek to solve the structure of the hijacked complex by X-ray crystallography. Critical to the success of this project is characterization of the disordered regions of recombinant Vif complexes using limited proteolysis in conjunction with mass-spectrometry. The information provided by these experiments will help design constructs for crystallization trials and allow determination of the structure of the Vif/host complexes, providing useful information for future antiretroviral therapies. A