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Substrate-inhibitor cooperative interactions with microbial dihydrofolate reductases
D P Baccanari1, R L Tansik, G H Hitchings
1Wellcome Research Laboratories, Research Triangle Park, North Carolina.
Advances in Enzyme Regulation
|January 1, 1987
Summary
This study investigates negative cooperativity in dihydrofolate reductase (DHFR) enzymes from Candida albicans and Saccharomyces cerevisiae, revealing distinct substrate binding mechanisms compared to E. coli DHFR. These findings impact understanding of antimicrobial drug resistance.
Area of Science:
- Biochemistry
- Enzymology
- Structural Biology
Background:
- Enzyme cooperativity, where substrate binding influences subsequent binding, is well-established.
- Positive cooperativity in dihydrofolate reductase (DHFR) enhances inhibitor binding, with variations across species.
- Negative cooperativity, a decrease in binding affinity upon forming a ternary complex, is less understood.
Purpose of the Study:
- To investigate negative cooperativity in DHFR from Candida albicans and Saccharomyces cerevisiae.
- To explore the relationship between inhibitor/NADPH cooperativity and trimethoprim (TMP) insensitivity in Neisseria gonorrhoeae.
- To compare structure-activity relationships of TMP analogs across different microbial DHFR enzymes.
Main Methods:
- Enzyme kinetics studies to assess substrate and inhibitor binding.
- Equilibrium binding assays to quantify cooperativity effects.
- Comparative analysis of DHFR enzymes from various microbial species.
Main Results:
- DHFR from C. albicans and S. cerevisiae exhibits strong negative cooperativity in substrate binding.
- Neisseria gonorrhoeae DHFR shows poor binary complex binding of TMP, with a 2,200-fold cooperative enhancement.
- Structure-activity relationships for TMP analogs differ significantly between E. coli and N. gonorrhoeae DHFR.
Conclusions:
- Negative cooperativity in yeast DHFR does not compromise intracellular activity due to physiologically relevant Michaelis constants.
- Understanding DHFR cooperativity mechanisms is crucial for developing effective antimicrobial strategies, particularly against resistant strains.
- Comparative enzymatic studies reveal species-specific drug interactions and resistance mechanisms.
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