Complementary Experimental Methods to Obtain Thermodynamic Parameters of Protein Ligand Systems
Abstract
:1. Introduction
2. Results and Discussion
2.1. EDTA–CaCl System
2.1.1. TDFRS Measurements
2.1.2. ITC Measurements
2.2. Protein–Ligand System
2.2.1. TDFRS Measurements
2.2.2. Isothermal Titration Calorimetry Measurements
2.2.3. Measurements with a Thermophoretic Microfluidic Cell
2.3. Validation of the Relation between Soret Coefficient and Gibb’s Free Energy
2.3.1. EDTA–CaCl System
2.3.2. Protein–Ligand System
3. Materials and Methods
3.1. Sample Preparation
3.1.1. EDTA–CaCl System
3.1.2. BCA-Ligand System
3.2. Methods
3.2.1. Thermal Diffusion Forced Rayleigh Scattering
3.2.2. Contrast Factor Measurement
3.2.3. Isothermal Titration Calorimetry
3.2.4. Thermophoretic Microfluidic Cells
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Data Availability Statement
Acknowledgments
Conflicts of Interest
Abbreviations
4FBS | 4-fluorobenzenesulfonamide |
BCA | Bovine Carbonic Anhydrase |
EDTA | ethylenediaminetetraacetic acide |
ITC | isothermal titration calorimetry |
MES | 2-(N-morpholino)ethanesulfonic acid |
PFBS | Pentafluorobenzenesulfonamide |
TDFRS | Thermal diffusion forced Rayleigh scattering |
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Temperature (°C) | K (nM) | H (kJ/mol) |
---|---|---|
20 | 510.0 ± 49.0 | −17.0 ± 0.3 |
25 | 623.0 ± 70.3 | −17.2 ± 0.8 |
30 | 699.0 ± 55.5 | −17.3 ± 0.7 |
35 | 852.0 ± 78.9 | −17.6 ± 0.5 |
40 | 1210.0 ± 123.0 | −17.8 ± 0.5 |
45 | 1570.0 ± 134.0 | −18.0 ± 0.8 |
System | K (nM) | H (kJ/mol) | G (kJ/mol) |
---|---|---|---|
BCA I–PFBS | 127.0 ± 47.2 | − 12.5 ± 0.8 | −37.4 ± 2.8 |
BCA I–4FBS | 325.0 ± 58.7 | −32.7 ± 0.4 | −37.5 ± 1.3 |
System | T (°C) | T (°C) | G (kJ/mol) | G (kJ/mol) |
---|---|---|---|---|
BCA I–PFBS | 30 | 20 | −40.5 ± 1.1 | −40.4 ± 1.3 |
BCA I–4FBS | 30 | 20 | −39.9 ± 3.9 | −38.2 ± 1.5 |
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Mohanakumar, S.; Lee, N.; Wiegand, S. Complementary Experimental Methods to Obtain Thermodynamic Parameters of Protein Ligand Systems. Int. J. Mol. Sci. 2022, 23, 14198. https://doi.org/10.3390/ijms232214198
Mohanakumar S, Lee N, Wiegand S. Complementary Experimental Methods to Obtain Thermodynamic Parameters of Protein Ligand Systems. International Journal of Molecular Sciences. 2022; 23(22):14198. https://doi.org/10.3390/ijms232214198
Chicago/Turabian StyleMohanakumar, Shilpa, Namkyu Lee, and Simone Wiegand. 2022. "Complementary Experimental Methods to Obtain Thermodynamic Parameters of Protein Ligand Systems" International Journal of Molecular Sciences 23, no. 22: 14198. https://doi.org/10.3390/ijms232214198