The proposed work focuses on basic structure/activity studies of two molybdenum-containing enzymes, xanthine oxidoreductase and carbon monoxide dehydrogenase. The first enzyme catalyzes the final two steps in purine metabolism in humans, and is also an important therapeutic target in the treatment of hyperuricemia and also in chemotherapeutic regimens. It has become the paradigm for mechanistic studies of the molybdenum hydroxylase family of enzymes. The second enzyme catalyzes the key step in carboxydotrophic growth on CO in certain bacteria and is responsible for removal of approximately 2 x 108 metric tons of CO from the environment each year. It is also a biological reaction of fundamental interest from a chemical standpoint: although CO dehydrogenase exhibits significant structural and sequence homologies to xanthine oxidoreductase, it is unique among the molybdenum hydroxylases in that its reaction does not involve C-H bond cleavage. The overall goal of the proposed work is to gain a more complete understanding of the mechanism of action of these two enzymes in the context of their structures, the guiding hypothesis being that enzyme function is dictated by the physical and electronic structure of the active site.
The Specific Aims i nclude rapid kinetic studies as well as spectroscopic work aimed at determining the electronic structures of intermediates identified. In the case of xanthine oxidoreductase, site-directed mutants targetting specific active site amino acid residues will be examined to evaluate their roles in catalysis. The overall goal is to compare and contrast the behavior of these two closely related enzymes so that the relationship of structure to function can be better understood.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
3R01GM075036-05S1
Application #
7892110
Study Section
Macromolecular Structure and Function A Study Section (MSFA)
Program Officer
Ikeda, Richard A
Project Start
2009-08-15
Project End
2010-06-30
Budget Start
2009-08-15
Budget End
2010-06-30
Support Year
5
Fiscal Year
2009
Total Cost
$163,091
Indirect Cost
Name
University of California Riverside
Department
Biochemistry
Type
Schools of Earth Sciences/Natur
DUNS #
627797426
City
Riverside
State
CA
Country
United States
Zip Code
92521
Lambeck, Iris C; Fischer-Schrader, Katrin; Niks, Dimitri et al. (2012) Molecular mechanism of 14-3-3 protein-mediated inhibition of plant nitrate reductase. J Biol Chem 287:4562-71
Cao, Hongnan; Hall, James; Hille, Russ (2011) X-ray crystal structure of arsenite-inhibited xanthine oxidase: ?-sulfido,?-oxo double bridge between molybdenum and arsenic in the active site. J Am Chem Soc 133:12414-7
Wilcoxen, Jarett; Zhang, Bo; Hille, Russ (2011) Reaction of the molybdenum- and copper-containing carbon monoxide dehydrogenase from Oligotropha carboxydovorans with quinones. Biochemistry 50:1910-6
Tejero, Jesus; Biswas, Ashis; Haque, Mohammad Mahfuzul et al. (2011) Mesohaem substitution reveals how haem electronic properties can influence the kinetic and catalytic parameters of neuronal NO synthase. Biochem J 433:163-74
Hille, Russ; Nishino, Takeshi; Bittner, Florian (2011) Molybdenum enzymes in higher organisms. Coord Chem Rev 255:1179-1205
Shanmugam, Muralidharan; Zhang, Bo; McNaughton, Rebecca L et al. (2010) The structure of formaldehyde-inhibited xanthine oxidase determined by 35 GHz 2H ENDOR spectroscopy. J Am Chem Soc 132:14015-7
Zhang, Bo; Hemann, Craig F; Hille, Russ (2010) Kinetic and spectroscopic studies of the molybdenum-copper CO dehydrogenase from Oligotropha carboxidovorans. J Biol Chem 285:12571-8
Pauff, James M; Cao, Hongnan; Hille, Russ (2009) Substrate Orientation and Catalysis at the Molybdenum Site in Xanthine Oxidase: CRYSTAL STRUCTURES IN COMPLEX WITH XANTHINE AND LUMAZINE. J Biol Chem 284:8760-7
Pauff, James M; Hille, Russ (2009) Inhibition studies of bovine xanthine oxidase by luteolin, silibinin, quercetin, and curcumin. J Nat Prod 72:725-31
Ilagan, Robielyn P; Tiso, Mauro; Konas, David W et al. (2008) Differences in a conformational equilibrium distinguish catalysis by the endothelial and neuronal nitric-oxide synthase flavoproteins. J Biol Chem 283:19603-15

Showing the most recent 10 out of 16 publications