The Oligomeric properties of the Adeno-Associated Virus Rep68 reflect its multifunctionality

Francisco Zarate-Perez, Jorge Mansilla-Soto, Martino Bardelli, John W Burgner, Maria Villamil-Jarauta, Demet Kekilli, Monserrat Samso, R Michael Linden, Carlos R Escalante

Research output: Contribution to journalArticlepeer-review

18 Citations (Scopus)


The Adeno-Associated Virus (AAV) encodes four regulatory proteins called Rep. The large AAV Rep proteins Rep68 and Rep78 proteins are essential factors required in almost every step of the viral life cycle. Structurally, they share two domains: a modified version of the AAA(+) domain that characterizes the SF3 family of helicases and an N-terminal domain that binds DNA specifically. The combination of these two domains imparts extraordinary multifunctionality to work as initiators of DNA replication and regulators of transcription, in addition to their essential role during site-specific integration. Although most members of the SF3 family form hexameric rings in vitro, the oligomeric nature of Rep68 is unclear due to its propensity to aggregate in solution. We report here a comprehensive study to determine the oligomeric character of Rep68 using a combination of methods that include sedimentation velocity ultracentrifugation, electron microscopy and hydrodynamic modeling. We have determined that residue Cys151 induces Rep68 to aggregate in vitro. We show that Rep68 displays a concentration dependent dynamic oligomeric behavior characterized by the presence of two populations: one with monomers and dimers in slow equilibrium and a second one consisting of a mixture of multiple ring structures of seven and eight members. The presence of either ATP or ADP induces formation of larger complexes formed by the stacking of multiple rings. Taken together, our results support the idea of a Rep68 molecule that exhibits the flexible oligomeric behavior needed to perform the wide range of functions occurring during the AAV viral life cycle.
Original languageEnglish
Pages (from-to)1232-1241
Number of pages10
JournalJournal of Virology
Issue number2
Early online date14 Nov 2012
Publication statusPublished - Jan 2013


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