Inositol hexakisphosphate (IP6) promotes HIV-1 assembly via its interaction with the immature Gag lattice, effectively enriching IP6 within virions. During particle maturation, the HIV-1 protease cleaves the Gag polyproteins comprising the immature Gag lattice, releasing IP6 from its original binding site and liberating the capsid (CA) domain of Gag. IP6 then promotes the assembly of mature CA protein into the capsid shell of the viral core, which is required for infection of new target cells. Recently, we reported HIV-1 Gag mutants that assemble virions independently of IP6. However, these mutants are non-infectious and unable to assemble stable capsids. Here, we identified a mutation in the C-terminus of CA - G225R - that restores capsid formation and infectivity to these IP6-packaging-deficient mutants. Furthermore, we show that G225R facilitates the