Because of their extreme importance and prevalence in nearly all biological processes, most industrial and academic laboratories are involved in some way in the characterization of ligand-macromolecule or macromolecule-macromolecule binding reactions. Up to the present, the measurement of equilibrium constants for the dissociation of such complexes has usually required the use of inconvenient, time-consuming methods based on partitioning (e.g., equilibrium dialysis or gel exclusion). Phase I involved the construction of a rapid titration calorimeter, utilizing much of the technology and hardware existing in MicroCal's scanning calorimeter, which will accurately determine both the dissociation constant, K, and heat of binding DeltaH, without the need of partitioning. The technique will be potentially applicable to all ligand-macromolecule and macromolecule-macromolecule interactions having a non-zero DeltaH and having K values in the range 10-2 to 10-8 M, which includes most biological binding processes. In many cases, a complete binding isotherm will be obtained in a single experiment lasting several minutes and consuming only nanomoles of biological macromolecule. Phase II will involve: 1.) Making further improvements in the instrument to increase overall performance and convenience. 2.) Construction of a modified injector/stirrer system to enable the instrument to be used for multiple-sample enzyme assays and for following reaction kinetics, in addition to characterizing binding reactions. 3.) Interfacing the calorimeter to an IBM-PC, and developing a large body of software for data collection and data analysis. 4.) Carrying out research on biological systems. 5.) Attempting to incorporate both scanning and titration capabilities in the same cell assembly.

Project Start
1986-09-29
Project End
1988-09-28
Budget Start
1987-09-29
Budget End
1988-09-28
Support Year
3
Fiscal Year
1987
Total Cost
Indirect Cost
Name
Microcal, Inc.
Department
Type
DUNS #
City
Northampton
State
MA
Country
United States
Zip Code
01060
Wiseman, T; Williston, S; Brandts, J F et al. (1989) Rapid measurement of binding constants and heats of binding using a new titration calorimeter. Anal Biochem 179:131-7