title: Crystallization and preliminary X-ray analysis of aspartate transcarbamoylase from the parasitic protist Trypanosoma cruzi.
abstract: Aspartate transcarbamoylase (ATCase), the second enzyme of the de novo pyrimidine-biosynthetic pathway, catalyzes the production of carbamoyl aspartate from carbamoyl phosphate and L-aspartate. In contrast to Escherichia coli ATCase and eukaryotic CAD multifunctional fusion enzymes, Trypanosoma cruzi ATCase lacks regulatory subunits and is not part of the multifunctional fusion enzyme. Recombinant T. cruzi ATCase expressed in E. coli was purified and crystallized in a ligand-free form and in a complex with carbamoyl phosphate at 277 K by the sitting-drop vapour-diffusion technique using polyethylene glycol 3350 as a precipitant. Ligand-free crystals (space group P1, unit-cell parameters a = 78.42, b = 79.28, c = 92.02 A, alpha = 69.56, beta = 82.90, gamma = 63.25 degrees) diffracted X-rays to 2.8 A resolution, while those cocrystallized with carbamoyl phosphate (space group P2(1), unit-cell parameters a = 88.41, b = 158.38, c = 89.00 A, beta = 119.66 degrees) diffracted to 1.6 A resolution. The presence of two homotrimers in the asymmetric unit (38 kDa x 6) gives V(M) values of 2.3 and 2.5 A(3) Da(-1) for the P1 and P2(1) crystal forms, respectively.
authors.affiliation: Department of Applied Biology, Graduate School of Science and Technology, Kyoto Institute of Technology, Kyoto, Japan.
authors.forename: Kazuaki
authors.initials: K
authors.lastname: Matoba
authors.forename: Takeshi
authors.initials: T
authors.lastname: Nara
authors.forename: Takashi
authors.initials: T
authors.lastname: Aoki
authors.forename: Teruki
authors.initials: T
authors.lastname: Honma
authors.forename: Akiko
authors.initials: A
authors.lastname: Tanaka
authors.forename: Masayuki
authors.initials: M
authors.lastname: Inoue
authors.forename: Shigeru
authors.initials: S
authors.lastname: Matsuoka
authors.forename: Daniel Ken
authors.initials: DK
authors.lastname: Inaoka
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Kita
authors.forename: Shigeharu
authors.initials: S
authors.lastname: Harada
mesh_terms: D000818:Animals; D001221:Aspartate Carbamoyltransferase / Q000737:chemistry*; D003460:Crystallization; D018360:Crystallography, X-Ray; D004591:Electrophoresis, Polyacrylamide Gel; D010271:Parasites / Q000201:enzymology*; D055672:Static Electricity; D014349:Trypanosoma cruzi / Q000201:enzymology*
chemical_list: D001221:Aspartate Carbamoyltransferase