Inhibitor binding to metal-substituted metalloenzyme: Sulfonamide affinity for carbonic anhydrase IX
Denis Baronas1, Birutė Knašienė2, Aurelija Mickevičiūtė1
1Department of Biothermodynamics and Drug Design, Institute of Biotechnology, Life Sciences Center, Vilnius University, Saulėtekio 7, Vilnius LT-10257, Lithuania.
Journal of Inorganic Biochemistry
|April 6, 2024
Summary
This study explores how different metal ions affect carbonic anhydrase (CA) enzyme activity and sulfonamide inhibitor binding. Understanding these metal-ion interactions is key to designing new drugs for metalloenzymes.
Area of Science:
- Biochemistry
- Enzymology
- Medicinal Chemistry
Background:
- Transition metal ions are essential cofactors for many proteins, including human carbonic anhydrase (CA), a zinc-dependent enzyme.
- Sulfonamide inhibitors target the zinc ion in the CA active site, but metal ions can be removed or substituted.
Purpose of the Study:
- To investigate the role of various divalent transition metal ions in the binding affinity of sulfonamide inhibitors to human carbonic anhydrase isozymes.
- To determine the intrinsic binding affinity, independent of pH-dependent protonation reactions, for metal-substituted CAs.
Main Methods:
- Fluorescence-based thermal shift assay was used to measure metal ion affinities to metal-free CA isozymes (CA I, II, IX).
- Metal-substituted CAs were prepared and used to determine sulfonamide compound binding.
- Binding-linked reactions were dissected to calculate intrinsic binding affinity.
Main Results:
- Sulfonamide inhibitor binding to metal-substituted CA exhibited a U-shaped pH dependence.
- For the cancer-associated isozyme CAIX, inhibitor binding affinity decreased in the order: Zn > Co > Hg > Cu > Cd > Mn > Ni.
- The energetic contribution of the inhibitor-metal coordination bond was quantified for tested metals.
Conclusions:
- The study elucidates the principles of how metal ions influence ligand affinity and selectivity in metalloenzymes.
- This understanding can guide the rational design of novel therapeutic agents targeting metalloenzymes.
Keywords:
Divalent metal ionFluorescent thermal shift assayMetal-free carbonic anhydrase (apoCA)Metal-substituted carbonic anhydraseMetalloenzymeSulfonamideMore Related Videos
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