ANALYSIS OF THE TERMINAL REPEAT BINDING ABILITIES OF MUTANT ADENOASSOCIATED VIRUS-REPLICATION PROTEINS

The adeno-associated virus (AAV) Rep78 and Rep68 proteins play essenti al roles in viral DNA replication, trans activation of viral gene expr ession, and suppression of oncogene-mediated cellular transformation. By using an extensive set of linker insertion and deletion mutations i n the replication gene, we mapped the regions of the Rep78 protein tha t mediate binding to the AAV origin of replication in vitro. Deletions that removed amino acid codons 25 to 62, 88 to 113, 125 to 256, and 3 46 to 400 abolished binding. Alterations in several other regions of t he protein affected the binding affinity of the mutant proteins. All o f the mutant proteins that support AAV DNA replication or p40 trans ac tivation bound to the terminal repeat sequence, thus verifying the imp ortance of binding for these functions. Several mutant rep genes that failed to suppress oncogene-mediated cellular transformation produced proteins that were capable of binding to the AAV terminal repeat seque nces.