Draft crystal structure of the vault shell at 9-A resolution.
Vaults are the largest known cytoplasmic ribonucleoprotein structures and may function in innate immunity. The vault shell self-assembles from 96 copies of major vault protein and encapsulates two other proteins and a small RNA. We crystallized rat liver vaults and several recombinant vaults, all among the largest non-icosahedral particles to have been crystallized. The best crystals thus far were formed from empty vaults built from a cysteine-tag construct of major vault protein (termed cpMVP vaults), diffracting to about 9-A resolution. The asymmetric unit contains a half vault of molecular mass 4.65 MDa. X-ray phasing was initiated by molecular replacement, using density from cryo-electron microscopy (cryo-EM). Phases were improved by density modification, including concentric 24- and 48-fold rotational symmetry averaging. From this, the continuous cryo-EM electron density separated into domain-like blocks. A draft atomic model of cpMVP was fit to this improved density from 15 domain models. Three domains were adapted from a nuclear magnetic resonance substructure. Nine domain models originated in ab initio tertiary structure prediction. Three C-terminal domains were built by fitting poly-alanine to the electron density. Locations of loops in this model provide sites to test vault functions and to exploit vaults as nanocapsules.
Anderson
Daniel H
DH
Howard Hughes Medical Institute, University of California Los Angeles, Los Angeles, California, United States of America.
Kickhoefer
Valerie A
VA
Sievers
Stuart A
SA
Rome
Leonard H
LH
Eisenberg
David
D
D000818:Animals; D001483:Base Sequence; D020285:Cryoelectron Microscopy; D003460:Crystallization; D008099:Liver / Q000737:chemistry*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry; D020394:Vault Ribonucleoprotein Particles / Q000737:chemistry*
D011994:Recombinant Proteins; D020394:Vault Ribonucleoprotein Particles