Thermodynamic characterization of human carbonic anhydrase VB stability and intrinsic binding of compounds

The thermodynamics of low molecular weight synthetic sulfonamide inhibitor binding to carbonic anhydrase (CA) VB was determined by the isothermal titration calorimetry (ITC) and the fluorescent thermal shift assay (FTSA). ITC provided the enthalpic and entropic contributions to the binding affinity...

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Bibliographic Details
Published in:Journal of thermal analysis and calorimetry 2016-03, Vol.123 (3), p.2191-2200
Main Authors: Kasiliauskaitė, Aistė, Časaitė, Vida, Juozapaitienė, Vaida, Zubrienė, Asta, Michailovienė, Vilma, Revuckienė, Jurgita, Baranauskienė, Lina, Meškys, Rolandas, Matulis, Daumantas
Format: Article
Language:English
Subjects:
Affinity
Analytical Chemistry
Assaying
Binding
Calorimetry
Carbon
Carbonic anhydrase
Chemical properties
Chemistry
Chemistry and Materials Science
Inhibitors
Inorganic Chemistry
Measurement Science and Instrumentation
Mitochondrial DNA
Obesity
Physical Chemistry
Physiological aspects
Polymer Sciences
Protein binding
Sulfonamides
Thermodynamics
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Summary:The thermodynamics of low molecular weight synthetic sulfonamide inhibitor binding to carbonic anhydrase (CA) VB was determined by the isothermal titration calorimetry (ITC) and the fluorescent thermal shift assay (FTSA). ITC provided the enthalpic and entropic contributions to the binding affinity of ethoxzolamide to CA VB. FTSA is a high-throughput assay that measures protein thermal stabilization by added ligands. FTSA enabled determination of extremely high affinity of several compounds binding to CA VB. CA VB is one of two isoforms that are expressed in mitochondria, participate in carbon metabolism and pH homeostasis and are implicated in diseases such as obesity. Therefore CA VB is a drug target. Here a series of para-substituted tetrafluoro benzenesulfonamides were investigated as high affinity inhibitors of CA VB. Thermodynamic equilibrium binding measurements such as ITC and FTSA provide only the observed parameters. Dissection of binding-linked reactions is necessary to obtain the intrinsic parameters that in turn could be correlated with the chemical structure of the inhibitors. Intrinsic dissociation constants of the inhibitors were estimated and they reached 1 pM, one of the strongest binding reactions observed between any protein–ligand binding.
ISSN:1388-6150
1588-2926
1572-8943
DOI:10.1007/s10973-015-5073-3