Studying enzyme kinetics through Isothermal Titration Calorimetry

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00:00:00 Welcome
00:01:23 Calorimetry- Basics
00:03:11 Isothermal Titration Calorimeter
00:04:55 Enzyme Kinetics
00:05:52 Enzyme Kinetics
00:07:41 Enzyme Kinetics and ITC
00:09:13 Determination of Enthalpy- M1
00:10:12 Experimental Considerations- M1
00:11:01 Calculating "True" Reaction Enthalpy
00:12:31 Determining dQ/dt
00:13:41 Experimental Considerations- M2
00:14:45 Determining dQ/dt
00:15:07 Continuous assay
00:15:40 Continuous assay
00:16:42 Experimental Considerations- Single Injection Method
00:18:01 Practical Considerations
00:19:40 Ni(ll)- Dependent Urease
00:20:41 Ni(ll)- Dependent Urease
00:21:10 Ni(ll)- Dependent Urease
00:22:12 Lactate Dehydrogenase
00:22:59 Lactate Dehydrogenase
00:23:22 Laccase
00:24:30 Enzyme Inhibition by ITC
00:26:05 Enzyme Inhibition by ITC
00:26:27 Urease Inhibition by Fluoride
00:28:23 LDH-A Inhibition by Oxamate
00:29:31 Trypsin Inhibition by Benzamidine
00:30:14 Conclusions
00:31:00 Acknowledgments
00:31:16 Thank you for your attentionAny questions?
00:45:48 Contact Information
Isothermal Titration Calorimetry (ITC) is a technique that measures the heat released or absorbed during a chemical reaction as an intrinsic probe to characterize any chemical process. In the present webinar, the experimental ITC-based methods to quantify kinetics and thermodynamics of enzymatic reactions are described.

ITC can concomitantly determine kinetics and thermodynamic parameters (kcat, KM, DH) of enzymatic reactions, not requiring any modification or labelling of the system under analysis, as heat changes spontaneously during enzymatic catalysis.

ITC can be performed in solution and needs little amount of material. These properties make this technique an invaluable, powerful and unique tool to extend the study of enzyme kinetics to several applications, such as drug discovery.