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Purification and characterization of fungal and mammalian phosphomannose isomerases
A E Proudfoot1, M A Payton, T N Wells
1Glaxo Institute for Molecular Biology, Geneva, Switzerland.
Abstract:
Phosphomannose isomerase (PMI) is essential for the production of yeast cell walls. An inhibitor which inhibits the fungal enzyme without altering the activity of the mammalian enzyme would be a potential fungicidal agent, increasingly important in view of the increasing mortality from visceral mycoses in immunosuppressed patients. We have purified human, porcine, and Candida albicans enzymes 29,000-fold to homogeneity, and characterized their physical properties, as well as their kinetic parameters, inhibition constants, and pH dependences. Surprisingly, in view of the large differences between Pseudomonas aerugenosa and Saccharomyces cerevisiae PMI, the human and C. albicans enzymes are almost identical. We suggest therefore that species-selective inhibition of the fungal rather than mammalian enzyme may require molecules which bind away from the substrate binding pocket of the enzyme.
Insights
Phosphomannose isomerase (PMI) is crucial for yeast cell walls. Targeting fungal PMI over human PMI could yield effective fungicidal agents against visceral mycoses.
Area of Science:
- Biochemistry
- Mycology
- Pharmacology
Background:
- Phosphomannose isomerase (PMI) is vital for yeast cell wall synthesis.
- Visceral mycoses pose a growing threat, especially in immunosuppressed individuals.
- Targeting fungal-specific enzymes is a key strategy for developing novel antifungal therapies.
Purpose of the Study:
- To purify and characterize human, porcine, and Candida albicans PMI.
- To investigate the kinetic and physical properties of these enzymes.
- To explore the potential for species-selective inhibition of fungal PMI.
Main Methods:
- Purification of human, porcine, and Candida albicans PMI to homogeneity (29,000-fold).
- Characterization of physical properties, kinetic parameters, and pH dependence.
- Determination of inhibition constants for potential inhibitors.
Main Results:
- Human and Candida albicans PMI enzymes exhibit remarkable similarity.
- Significant differences exist between Pseudomonas aeruginosa and Saccharomyces cerevisiae PMI.
- Species-selective inhibition may necessitate targeting sites distinct from the substrate-binding pocket.
Conclusions:
- Human and fungal PMI share high structural and functional similarity.
- Developing selective inhibitors requires understanding enzyme variations.
- Allosteric inhibition or targeting unique structural features may be necessary for selective antifungal agents.