Structural studies of AAV2 Rep68 reveal a partially structured linker and compact domain conformation

Faik N Musayev, Francisco Zarate-Perez, Martino Raul Bardelli, Clayton M Bishop, Emil F Saniev, R Michael Linden, Els Henckaerts, Carlos R Escalante

Research output: Contribution to journalArticlepeer-review

8 Citations (Scopus)

Abstract

Adeno-associated virus (AAV) non-structural proteins Rep78 and Rep68 carry out all DNA transactions that regulate the AAV life cycle. They share two multifunctional domains: an N-terminal origin binding/nicking domain (OBD) from the HUH superfamily and a SF3 helicase domain. A short linker of ~ 20 amino acids that is critical for oligomerization and function connects the two domains. Although X-ray structures of the AAV5 OBD and AAV2 helicase domains have been determined, information about the full-length protein and linker conformation is not known. This paper presents the solution structure of AAV2 Rep68 using small-angle x-ray scattering (SAXS). We first determined the X-ray structures of the minimal AAV2 Rep68 OBD and of the OBD with the linker region. These X-ray structures reveal novel features that include a long C-terminal α-helix that protrudes from the core of the protein at a 45° angle and a partially structured linker. SAXS studies corroborate that the linker is not extended and we show that a proline residue in the linker is critical for Rep68 oligomerization and function. SAXS-based rigid body modeling of Rep68 confirms these observations showing a compact arrangement of the two domains acquiring a conformation that positions key residues in all domains on one face of the protein and poised to interact with DNA.
Original languageEnglish
JournalBiochemistry
DOIs
Publication statusE-pub ahead of print - 28 Aug 2015

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