Phenylbutazone radicals inactivate creatine kinase.

Area of Science:

• Biochemistry
• Pharmacology
• Toxicology

Background:

• Phenylbutazone (PB) is a drug used for rheumatic and arthritic diseases.
• Tissue damage is a potential side effect of PB.
• Creatine kinase (CK) is a sensitive enzyme marker for cellular damage.

Purpose of the Study:

• To investigate the mechanism of tissue damage induced by phenylbutazone (PB).
• To identify the role of creatine kinase (CK) as a marker for PB-induced side effects.
• To elucidate the interaction between PB, CK, and reactive oxygen species.

Main Methods:

• Enzyme inactivation assays using CK and other enzymes.
• Detection of PB carbon-centered radicals using electron spin resonance (ESR).
• Use of spin trap agents to identify radical intermediates.

Main Results:

• PB inactivated CK in the presence of horseradish peroxidase and hydrogen peroxide (HRP-H(2)O2).
• PB carbon-centered radicals were generated and detected during the PB-HRP-H(2)O2 interaction.
• CK reduced ESR signals of PB radicals, and spin traps prevented CK inactivation, indicating radical-mediated damage.
• Loss of sulfhydryl and tryptophan residues in CK, suggesting oxidation of cysteine and tryptophan by PB radicals.
• Other sulfhydryl-containing enzymes were also inactivated, but lactate dehydrogenase was not.

Conclusions:

• Creatine kinase (CK) is inactivated by phenylbutazone (PB) through a mechanism involving PB carbon-centered radicals.
• The oxidation of cysteine and tryptophan residues in CK by these radicals contributes to enzyme inactivation.
• Sulfhydryl-dependent enzymes are particularly susceptible to PB-induced radical damage.
• Inhibition of sulfhydryl enzymes may underlie some of the adverse effects associated with PB treatment.