title: Crystallization of hamster dihydroorotase: involvement of a disulfide-linked tetrameric form.
abstract: Dihydroorotase (DHOase) catalyses the formation of L-dihydroorotate (DHO) in the de novo pyrimidine biosynthetic pathway. The type I DHOase domain from hamster forms part of the trifunctional enzyme CAD. The hamster DHOase domain has been cloned and expressed in Escherichia coli. Solutions of the homodimeric protein convert to a homotetrameric species when incubated at ambient temperature. Formation of the tetrameric species is mediated via disulfide linkages between single free cysteine residues on the surface of each monomer. This process is also observed under conditions used for crystallization of the hamster DHOase domain; crystals composed exclusively of the tetrameric species grow from solutions containing as little as 10% tetramer. The crystallization of pure tetrameric DHOase results in two crystal forms: form I, with space group C222(1) and unit-cell parameters a = 127.1, b = 603.5, c = 144.7 A, and form II, with space group P2(1) and unit-cell parameters a = 260.5, b = 148.2, c = 308.0 A, beta = 102.2 degrees. Data have been recorded to 4.3 and 4.0 A resolution, respectively.
authors.affiliation: School of Molecular and Microbial Biosciences, University of Sydney, Sydney, NSW 2006, Australia. m.maher@mmb.usyd.edu.au
authors.forename: Megan J
authors.initials: MJ
authors.lastname: Maher
authors.forename: Danny T C
authors.initials: DT
authors.lastname: Huang
authors.forename: J Mitchell
authors.initials: JM
authors.lastname: Guss
authors.forename: Charles A
authors.initials: CA
authors.lastname: Collyer
authors.forename: Richard I
authors.initials: RI
authors.lastname: Christopherson
mesh_terms: D000818:Animals; D003001:Cloning, Molecular; D006224:Cricetinae; D003460:Crystallization; D018360:Crystallography, X-Ray; D003545:Cysteine / Q000737:chemistry; D004080:Dihydroorotase / Q000737:chemistry* / Q000235:genetics; D004220:Disulfides / Q000737:chemistry*; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics
chemical_list: D004220:Disulfides; D011994:Recombinant Proteins; D004080:Dihydroorotase; D003545:Cysteine