Functional domain organization of human APOBEC3G

Area of Science:

• Virology
• Molecular Biology
• Genetics

Background:

• Human APOBEC3 proteins are key antiviral factors, with varying inhibitory effects on HIV-1.
• APOBEC3 proteins exist as single or dual cytidine deaminase domain (CDA) forms.
• APOBEC3G (A3G) with two CDAs potently inhibits HIV-1, while APOBEC3A (A3A) with one CDA is a weak inhibitor despite its deaminase activity.

Purpose of the Study:

• To investigate the functional roles of CDA domains in APOBEC3 protein activity against HIV-1.
• To determine the regions responsible for Gag binding and virion packaging in A3G.
• To assess if conferring Gag-binding and packaging ability to A3A enhances its HIV-1 inhibitory function.

Main Methods:

• Site-directed mutagenesis of APOBEC3A (A3A) to mimic the N-terminal domain of APOBEC3G (A3G).
• Assays to evaluate Gag binding and virion incorporation of modified A3A.
• Infectivity assays of Vif-deficient HIV-1 (HIV-1ΔVif) in the presence of A3A mutants.

Main Results:

• The N-terminal half of A3G contains the determinants for Gag binding and virion packaging.
• A modified A3A protein, incorporating three amino acids from A3G's N-terminal domain, efficiently bound Gag and was packaged into HIV-1 virions.
• Despite enhanced virion incorporation, the modified A3A remained a weak inhibitor of HIV-1ΔVif infectivity.

Conclusions:

• Virion packaging and Gag binding activities of APOBEC3 proteins are localized to the N-terminal CDA domain.
• Efficient inhibition of HIV-1 infectivity by APOBEC3 proteins requires both Gag-binding/packaging and cytidine deaminase activities.
• The segregation of these activities into two tandem CDA domains, as seen in A3G, is crucial for potent retroviral inhibition.