Ethanenitronate is a peroxide-dependent suicide substrate for catalase

Area of Science:

• Biochemistry
• Enzymology
• Chemical Kinetics

Background:

• Bovine liver catalase (E) catalyzes the decomposition of hydrogen peroxide.
• Enzyme inactivation mechanisms are crucial for understanding enzyme function and regulation.
• Suicide substrates offer a unique approach to enzyme inhibition studies.

Purpose of the Study:

• To investigate the mechanism of ethanenitronate-induced inactivation of bovine liver catalase.
• To elucidate the role of radical intermediates in the inactivation process.
• To characterize the modified enzyme species formed during inactivation.

Main Methods:

• Enzyme kinetics assays using H2O2.
• Spectroscopic analysis of enzyme intermediates.
• Use of radiolabeled ethanenitronate ([1-14C]) to identify modification sites.

Main Results:

• Ethanenitronate is a H2O2-dependent suicide substrate for bovine liver catalase.
• Inactivation proceeds via a free radical intermediate (EN.) which primarily targets Compound II.
• A modified enzyme species (Em) is formed, retaining 7% of the original catalytic activity and covalently bound 14C-ferriheme.

Conclusions:

• The inactivation mechanism involves the trapping of the EN. radical by the catalase apoenzyme, leading to irreversible formation of Em.
• This mechanism is analogous to that observed for horseradish peroxidase, with differences in the trapping site.
• The labeled residue in Em is likely located near the Compound II reduction site.