INVESTIGATION OF POTENTIAL NOVEL DOMAIN WITHIN A3G: AN ANTI-HIV PROTEIN

Background: APOBEC3G (A3G) is a human antiviral protein that functions as an important cellular defense against HIV-1 infection. It catalyzes cytidine deamination ultimately resulting in mutagenesis of the HIV-1 genome during the essential process of reverse transcription. Unexpectedly A3G has also been shown to exhibit antiviral function in the absence of this mutagenic function. A fundamental molecular understanding of this deamination-independent mechanism awaits characterization. Methods: To delineate essential regions of A3G that contribute to its antiviral capabilities, a library of mutant A3G expression constructs were analyzed for their ability to suppress HIV-1 infection. 293T cells were co-transfected with HIV-1 provirus and the A3G mutant construct of interest. Resulting virus particles were isolated, quantitated and then assessed for infectivity in a single-round infectivity assay utilizing reporter cells. Essential virion packaging of the A3G mutant proteins was also examined via western blotting and the enzymatic (mutagenic) potential of the mutants was also quantified in an E. colibased system. Results: Mutants no longer capable of viral restriction, but continued to be catalytically active primarily exhibited mutations in two N-terminal regions of the A3G protein. These unique domains are implicated in the playing important roles in the deaminase-independent antiviral mechanism of A3G. Conclusion: These novel domains provide a platform from which to characterize the deaminase-independent mechanism of A3G that is currently poorly understood. Further description of the mechanistic role of these regions may aid in defining useful targets for antiviral therapeutics.