The reaction of ozone with glyceraldehyde-3-phosphate dehydrogenase
Inactivation of glyceraldehyde-3-phosphate dehydrogenase (GPDH) by ozone can be correlated with oxidation of the active-site -SH residue. Oxidation of peripheral -SH groups, and tryptophan, methionine, and histidine residues occurs concomitantly, but loss of activity depends solely on active-site oxidation. Inactivation is only slightly reversible by dithiothreitol. Kinetic studies show that inhibition of GPDH by ozone mimics noncompetitive inhibition and is characterized as irreversible enzyme inactivation. Analysis of products resulting from ozone oxidation of glutathione suggests that cysteic acid is the product of protein-SH oxidation. Despite oxidation of the active-site -SH, no significant decrease in the Racker band absorbance occurs. This is explained by the appearance of a new chromophore in this region of the absorbance spectrum. Increased absorbance at 322 nm following ozone treatment indicates that tryptophan is converted quantitatively to N-formylkynurenine. When the active-site -SH is reversibly blocked by tetrathionate, enzyme activity is completely recoverable following reaction of the derivatized enzyme with a 1.3X excess of ozone over enzyme monomer. Activity is fully recovered despite the oxidation of peripheral -SH, tryptophan, and histidine residues. Circular dichroism spectra of ozone-treated enzyme show that reaction of GPDH with up to a threefold excess of ozone over enzyme monomer results in no significant disruption of protein secondary structure. Spectra in the near-uv show distinct changes that reflect tryptophan oxidation.
- Research Organization:
- Department of Biochemistry, University of California, Riverside
- OSTI ID:
- 6963206
- Journal Information:
- Arch. Biochem. Biophys.; (United States), Vol. 229:1
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
OXIDOREDUCTASES
INHIBITION
OZONE
BIOLOGICAL EFFECTS
AMINO ACIDS
BIOCHEMISTRY
ENZYME ACTIVITY
OXIDATION
PROTEINS
CARBOXYLIC ACIDS
CHEMICAL REACTIONS
CHEMISTRY
ENZYMES
ORGANIC ACIDS
ORGANIC COMPOUNDS
560301* - Chemicals Metabolism & Toxicology- Cells- (-1987)