title: A C-RING-like domain participates in protein self-assembly and mineral nucleation.
abstract: AP7 is a nacre-associated protein of the mollusk shell that forms supramolecular assemblies that nucleate single-crystal aragonite in vitro. AP7 possesses two major sequence regions: a random coil 30-amino acid N-terminal domain (AP7N) and a partially disordered 36-amino acid C-terminal domain (AP7C) that exhibits imperfect sequence homology to the C subclass of the intracellular RING domain family. We report here new findings that implicate the C-RING domain in AP7-mediated supramolecular assembly and single-crystal mineral formation. AP7 protein spontaneously self-assembles over a pH range of 4-9 and is monomeric at pH >9.5. AP7N and AP7C both oligomerize over the pH range of 4-9, with the AP7C sequence closely resembling AP7 in terms of particle morphology and size. In vitro mineralization experiments demonstrate that both AP7N and AP7C form supramolecular assemblies that nucleate single-crystal calcium carbonates. Comparison of previously published nuclear magnetic resonance-based structures of AP7C and AP7N reveals the significant presence of complementary anionic-cationic electrostatic molecular surfaces on AP7C that are not found on AP7N, and this may explain the noted discrepancies between the two domains in terms of self-assembly and single-crystal nucleation. We conclude that the C-RING-like sequence is an important site for AP7 self-association and mineral nucleation, and this represents the first known instance of a RING-like sequence performing these functions within an extracellular protein.
authors.affiliation: Laboratory for Chemical Physics, New York University, 345 East 24th Street, New York, New York 10010, USA.
authors.forename: Fairland F
authors.initials: FF
authors.lastname: Amos
authors.forename: Moise
authors.initials: M
authors.lastname: Ndao
authors.forename: Christopher B
authors.initials: CB
authors.lastname: Ponce
authors.forename: John Spencer
authors.initials: JS
authors.lastname: Evans
mesh_terms: D000818:Animals; D000838:Anions; D002119:Calcium Carbonate / Q000737:chemistry; D002352:Carrier Proteins / Q000737:chemistry*; D002412:Cations; D006863:Hydrogen-Ion Concentration; D008027:Light; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D046529:Microscopy, Electron, Transmission / Q000379:methods; D008903:Minerals / Q000737:chemistry; D008974:Mollusca; D060734:Nacre / Q000737:chemistry*; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry; D012542:Scattering, Radiation; D055672:Static Electricity
chemical_list: D000838:Anions; D002352:Carrier Proteins; D002412:Cations; D008903:Minerals; D060734:Nacre; D011506:Proteins; D002119:Calcium Carbonate