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Project

Physical and Biophysical Chemistry Division (I)

 

Number: 2001-030-1-100

Title: Recommendations on the measurement and analysis of results obtained on biological substances with isothermal titration calorimetry

Task Group
Chairmen
: Frederick P. Schwarz and Hans-Jurgen Hinz

Completion Date: 2008 - project completed

Objective:
Recommendations for isothermal titration calorimetry measurement procedures on biological substances, calibration procedures, and procedures for testing the performance of isothermal titration calorimeters (ITCs) with a biological test solution are described. Recommendations for the analysis and reporting of the results will be presented to facilitate universal comparability of ITC data from different laboratories.

Description:
Isothermal titration calorimeters (ITCs) have been widely used in the biotechnological and pharmaceutical industry and in academia to determine the thermodynamics of binding reactions of biological substances, including DNA-DNA, DNA-RNA, protein-protein, protein-ligand, and DNA-intercalating drug interactions. ITCs monitor the power exchanged between a reference vessel and a solution vessel as aliquots of a titrant solution are injected into the solution vessel, isothermally. The enthalpy change, binding affinity, and the stoichiometry for the binding reaction are obtained from an analysis of the power exchanged per each injection and the total concentrations of the reactants in the solution vessel. In addition, the temperature dependence of these quantities can be determined in terms of a heat capacity change for the binding reaction. Thus, the thermodynamics of the binding reaction can be completely characterized over a wide temperature range.

Published results sometimes lack a systematic reporting and analysis of the data and often employ a variety of notation inconsistent with IUPAC recommendations. An important aspect of this project is the fabrication of a test solution for determination of ITC binding data. ITC test solutions will be distributed to members of the working party to run in their ITCs so that a consensus set of binding data can be developed on a particular system under a specified set of operating conditions. This consensus data can then be used to determine if a particular ITC is being operated properly. This is particularly important for new and in-experienced researchers employing ITC for the first time. The results will also be shared with the MIRG committee of the Association of Biomolecular Research Facilities (ABRF). MIRG is involved in evaluating how well binding affinities determined by ITC, Biacore SPR, and Ultra-analytical centrifugation are in agreement.

Progress:
June 2005 - Measurements are complete for a working standard NAD/NADH binding to a protein, lactate-dehydrogenase for checking the performance of isothermal titration calorimeters. The first draft of the report is will be complete by December 2005. The ‘round-robin’ ITC results from 12 laboratories on the binding of 4 carboxybenzene sulfonamide to carbonic anhydrase are complete and they are being evaluated for inclusion in the IUPAC Recommendations.

Sep 2008 - Project completed - IUPAC Technical Report published in Pure Appl. Chem., 2008, Vol. 80, No. 9, pp. 2025-2040

Last update: 19 September 2008

 

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Page last modified 19 September 2008.
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