title: Disulfide bond stabilization of the hexameric capsomer of human immunodeficiency virus.
abstract: The human immunodeficiency virus type 1 capsid is modeled as a fullerene cone that is composed of approximately 250 hexamers and 12 pentamers of the viral CA protein. Structures of CA hexamers have been difficult to obtain because the hexamer-stabilizing interactions are inherently weak, and CA tends to spontaneously assemble into capsid-like particles. Here, we describe a two-step biochemical strategy to obtain soluble CA hexamers for crystallization. First, the hexamer was stabilized by engineering disulfide cross-links (either A14C/E45C or A42C/T54C) between the N-terminal domains of adjacent subunits. Second, the cross-linked hexamers were prevented from polymerizing further into hyperstable capsid-like structures by mutations (W184A and M185A) that interfered with dimeric association between the C-terminal domains that link adjacent hexamers. The structures of two different cross-linked CA hexamers were nearly identical, and we combined the non-mutated portions of the structures to generate an atomic resolution model for the native hexamer. This hybrid approach for structure determination should be applicable to other viral capsomers and protein-protein complexes in general.
authors.affiliation: Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville, VA 22908, USA.
authors.forename: Owen
authors.initials: O
authors.lastname: Pornillos
authors.forename: Barbie K
authors.initials: BK
authors.lastname: Ganser-Pornillos
authors.forename: Sankaran
authors.initials: S
authors.lastname: Banumathi
authors.forename: Yuanzi
authors.initials: Y
authors.lastname: Hua
authors.forename: Mark
authors.initials: M
authors.lastname: Yeager
mesh_terms: D036022:Capsid Proteins / Q000737:chemistry / Q000378:metabolism*; D020285:Cryoelectron Microscopy; D003460:Crystallization; D018360:Crystallography, X-Ray; D004220:Disulfides / Q000737:chemistry / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D015497:HIV-1 / Q000378:metabolism*; D008958:Models, Molecular; D011487:Protein Conformation
chemical_list: D036022:Capsid Proteins; D004220:Disulfides