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Evidence for two distinct active sites on aldehyde dehydrogenase
The Journal of Biological Chemistry
|January 25, 1988
Summary
Horse liver aldehyde dehydrogenase exhibits distinct active sites for ester hydrolysis and aldehyde oxidation. Cysteine 49 is crucial for aldehyde oxidation, while both sites participate in ester hydrolysis.
Area of Science:
- Biochemistry
- Enzymology
Background:
- Aldehyde dehydrogenase (ALDH) possesses both esterase and dehydrogenase activities.
- The precise localization of these activities within the enzyme's active site(s) remains debated.
Purpose of the Study:
- To investigate whether the esterase and dehydrogenase activities of aldehyde dehydrogenase occur at the same active site.
- To elucidate the roles of specific cysteine residues in enzyme function.
Main Methods:
- Differential protection of enzyme activities against inactivation using substrate analogs.
- Chemical modification of cysteine residues using N-ethylmaleimide.
- Enzyme kinetics studies to assess inhibition patterns.
Main Results:
- Enzyme activities showed differential protection against inactivation.
- Aldehydes were poor inhibitors of esterase activity, while esters effectively inhibited dehydrogenase activity.
- Modification of Cys49 did not inhibit esterase activity, but modification of Cys162 was required for esterase inhibition, suggesting distinct roles.
Conclusions:
- Horse liver aldehyde dehydrogenase appears to possess two separate active sites per subunit.
- Ester hydrolysis occurs at both sites, whereas aldehyde oxidation is localized to the site involving Cys49.