Abstract
This chapter describes how to collect Michaelis-Menten kinetic data on an enzyme-catalyzed reaction with the isothermal titration calorimetry (ITC) and the single-injection method. ITC measures the heat rate which is directly proportional to the reaction rate. The enthalpy change (ΔrH), Km, kcat, and vmax are determined in a single assay that does not require labeling or immobilization.
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References
Todd M, Gomez J (2001) Enzyme kinetics determined using calorimetry: a general assay for enzyme activity? Anal Biochem 296:179–187
Morin PE, Friere E (1991) Direct calorimetric analysis of the enzymatic activity of yeast cytochrome c oxidase. Biochemist 30:8494–8500
Olsen S (2006) Applications of isothermal titration calorimetry to measure enzyme kinetics and activity in complex solutions. Thermochim Acta 448:12–18
Choma CT (2006). Characterizing enzyme kinetics by ITC. Applications Note M125, TA Instruments. http://www.tainstruments.com. Accessed 06 Sept 2012
Bianconi ML (2003) Calorimetric determination of thermodynamic parameters of reaction reveals different enthalpic compensations of the yeast hexokinase isozymes. J Biol Chem 278(21):18709–18713
Transtrum M, Hansen L, Quinn C (2015) Enzyme kinetics determined by single-injection isothermal titration calorimetry. Methods 76:194–200
Transtrum M, Hansen L, Quinn C, Demarse N (2016) Enzyme-catalyzed and binding reaction kinetics determined by titration calorimetry. Biochim Biophys Acta 1860:957–966
Hansen CW, Hansen LD, Nicholson AD et al (2011) Correction for instrument time constant and baseline in determination of chemical kinetics. International J Chem Kinetics 43(2):53–61
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Quinn, C.F., Hansen, L.D. (2019). Enzyme Kinetics Determined by Single-Injection Isothermal Titration Calorimetry. In: Ennifar, E. (eds) Microcalorimetry of Biological Molecules. Methods in Molecular Biology, vol 1964. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-9179-2_17
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DOI: https://doi.org/10.1007/978-1-4939-9179-2_17
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Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-9178-5
Online ISBN: 978-1-4939-9179-2
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