COORDINATED DISINTEGRATION REACTIONS MEDIATED BY MOLONEY MURINE LEUKEMIA-VIRUS INTEGRASE

The protein-DNA and protein-protein interactions important for functio n of the integrase (IN) protein of Moloney murine leukemia virus (M-Mu LV) were investigated by using a coordinated-disintegration assay. A p anel of M-MuLV IN mutants and substrate alterations highlighted distin ctions between the intermolecular and intramolecular reactions of coor dinated disintegration. Mispairing of the crossbone single-strand regi on and altered long terminal repeat (LTR) positioning affected the int ermolecular, but not the intramolecular, reactions of coordinated disi ntegration. Partial components of the crossbone substrate were coordin ated by M-MuLV IN, indicating a reliance on both LTR and target DNA de terminants for substrate assembly. The intramolecular reaction was dep endent on the presence of either the HHCC domain or a crossbone LTR 5' single-stranded tail. An M-MuLV IN mutant without the HHCC domain (N Delta 105) catalyzed reduced levels of double disintegration but not s ingle disintegration. A separately purified HHCC domain protein (C Del ta 232) stimulated double disintegration mediated by N Delta 105, sugg esting a role of the N-terminal HHCC domain in stable IN-IN and IN-DNA interactions. Significantly, crossbone substrates lacking the LTR 5' tails were not recognized by the fingerless N Delta 105 protein. Colle ctively, these data suggest similar roles of the HHCC domain and 5' LT R tail in substrate recognition and modulation of IN activity.