Substrate Orientation and Catalysis at the Molybdenum Site in Xanthine Oxidase: CRYSTAL STRUCTURES IN COMPLEX WITH XANTHINE AND LUMAZINE.
Xanthine oxidoreductase is a ubiquitous cytoplasmic protein that catalyzes the final two steps in purine catabolism. We have previously investigated the catalytic mechanism of the enzyme by rapid reaction kinetics and x-ray crystallography using the poor substrate 2-hydroxy-6-methylpurine, focusing our attention on the orientation of substrate in the active site and the role of Arg-880 in catalysis. Here we report additional crystal structures of as-isolated, functional xanthine oxidase in the course of reaction with the pterin substrate lumazine at 2.2 A resolution and of the nonfunctional desulfo form of the enzyme in complex with xanthine at 2.6 A resolution. In both cases the orientation of substrate is such that the pyrimidine subnucleus is oriented opposite to that seen with the slow substrate 2-hydroxy-6-methylpurine. The mechanistic implications as to how the ensemble of active site functional groups in the active site work to accelerate reaction rate are discussed.
Pauff
James M
JM
Department of Biochemistry, University of California, Riverside, California 92521 and the Medical Scientist Program, The Ohio State University, Columbus, Ohio 43210, USA.
Cao
Hongnan
H
Hille
Russ
R
D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry; D002384:Catalysis; D020134:Catalytic Domain / Q000502:physiology; D002417:Cattle; D018360:Crystallography, X-Ray; D005260:Female; D008892:Milk / Q000201:enzymology*; D008982:Molybdenum / Q000737:chemistry*; D017434:Protein Structure, Tertiary / Q000502:physiology; D011621:Pteridines / Q000737:chemistry*; D016354:Rhodobacter capsulatus / Q000201:enzymology; D019820:Xanthine / Q000737:chemistry*; D014969:Xanthine Oxidase / Q000737:chemistry* / Q000302:isolation & purification
D001426:Bacterial Proteins; D011621:Pteridines; D019820:Xanthine; C008297:lumazine; D008982:Molybdenum; D014969:Xanthine Oxidase