A preliminary neutron diffraction study of rasburicase, a recombinant urate oxidase enzyme, complexed with 8-azaxanthin.
Crystallization and preliminary neutron diffraction measurements of rasburicase, a recombinant urate oxidase enzyme expressed by a genetically modified Saccharomyces cerevisiae strain, complexed with a purine-type inhibitor (8-azaxanthin) are reported. Neutron Laue diffraction data were collected to 2.1 A resolution using the LADI instrument from a crystal (grown in D2O) with volume 1.8 mm3. The aim of this neutron diffraction study is to determine the protonation states of the inhibitor and residues within the active site. This will lead to improved comprehension of the enzymatic mechanism of this important enzyme, which is used as a protein drug to reduce toxic uric acid accumulation during chemotherapy. This paper illustrates the high quality of the neutron diffraction data collected, which are suitable for high-resolution structural analysis. In comparison with other neutron protein crystallography studies to date in which a hydrogenated protein has been used, the volume of the crystal was relatively small and yet the data still extend to high resolution. Furthermore, urate oxidase has one of the largest primitive unit-cell volumes (space group I222, unit-cell parameters a = 80, b = 96, c = 106 A) and molecular weights (135 kDa for the homotetramer) so far successfully studied with neutrons.
Budayova-Spano
Monika
M
European Molecular Biology Laboratory Grenoble Outstation, 38042 Grenoble, France. spano@embl-grenoble.fr
Bonneté
Françoise
F
Ferté
Natalie
N
El Hajji
Mohamed
M
Meilleur
Flora
F
Blakeley
Matthew Paul
MP
Castro
Bertrand
B
D001665:Binding Sites; D003460:Crystallization / Q000379:methods; D017666:Deuterium Oxide; D033363:Neutron Diffraction / Q000379:methods; D011994:Recombinant Proteins / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D014503:Urate Oxidase / Q000737:chemistry*; D014970:Xanthines / Q000737:chemistry*
D011994:Recombinant Proteins; D014970:Xanthines; C469709:rasburicase; C018221:8-azaxanthine; D014503:Urate Oxidase; D017666:Deuterium Oxide