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Updated: Feb 16, 2026
Fast and Specific Assessment of the Halogenating Peroxidase Activity in Leukocyte-enriched Blood Samples
Published on: July 28, 2016
A multi-substrate assay for finding physiologically effective inhibitors of myeloperoxidase
Louisa V Forbes1, Anthony J Kettle1
1Centre for Free Radical Research, Department of Pathology, University of Otago Christchurch, P.O. Box 4546, Christchurch, New Zealand.
Abstract:
Myeloperoxidase, an abundant neutrophil enzyme, promotes oxidative damage during inflammation by generating hypohalous acids and free radicals. Currently, there are no selective drugs to inhibit its adverse activity. This short-coming is partly due to the lack of screening assays that mimic the complex enzymatic activities of myeloperoxidase in vivo. We have developed an assay for myeloperoxidase activity that includes its major physiological substrates - chloride, thiocyanate, tyrosine, and urate. The multi-substrate assay monitors bleaching of 5-thio-2-nitrobenzoic acid and measures total oxidant production when hydrogen peroxide activates the enzyme. Known suicide inhibitors and tight-binders tested positive in the assay, whereas compounds that merely convert myeloperoxidase to reducible enzyme intermediates were poor inhibitors. The new assay revealed that some aromatic compounds, including tryptamine, inhibit myeloperoxidase by binding reversibly to the enzyme. Our multi-substrate assay is selective for physiologically relevant inhibitors and has potential for identifying new classes of myeloperoxidase inhibitors.
Insights
A new assay detects myeloperoxidase (MPO) inhibitors by mimicking its in vivo activity. This tool identifies selective MPO inhibitors, including novel aromatic compounds, crucial for treating inflammatory diseases.
Area of Science:
- Biochemistry
- Enzymology
- Pharmacology
Background:
- Myeloperoxidase (MPO) is a neutrophil enzyme driving oxidative damage in inflammation.
- Lack of selective MPO inhibitors stems from inadequate in vivo mimicking assays.
- Existing assays fail to capture MPO's complex enzymatic functions.
Purpose of the Study:
- Develop a novel screening assay for myeloperoxidase (MPO) activity.
- Incorporate MPO's major physiological substrates (chloride, thiocyanate, tyrosine, urate).
- Identify selective MPO inhibitors with therapeutic potential.
Main Methods:
- Created a multi-substrate assay for MPO activity.
- Monitored 5-thio-2-nitrobenzoic acid bleaching.
- Measured total oxidant production upon enzyme activation by hydrogen peroxide.
Main Results:
- The assay successfully identified known MPO suicide inhibitors and tight-binders.
- Compounds causing only reversible enzyme intermediates showed weak inhibition.
- Discovered novel reversible MPO inhibitors, including aromatic compounds like tryptamine.
Conclusions:
- The developed multi-substrate assay selectively identifies physiologically relevant MPO inhibitors.
- This assay is a valuable tool for discovering new classes of MPO-targeting drugs.
- Potential for developing treatments for inflammatory conditions driven by MPO.
