affinimeter & isothermal titration calorimetry
TRANSCRIPT
for the complete Thermodynamic and Kinetic Characterization of molecular interactions
(1) AFFINImeter Scientific & Development team, Software 4 Science Developments, S. L. Ed. Emprendia. �(2) Dept. of Applied Physics, Fac. of Physics, USC, Campus Vida, Santiago de Compostela, A Coruña 15782, Spain.
(3) Biophysics & Structural Biology team, IBMC, UPR9002 du CNRS 15 rue René Descartes, 67084 Strasbourg, France. Website: www.affinimeter.com; E-mail: [email protected]
AFFINImeter & Isothermal Titration Calorimetry
1.- GET THERMODYNAMIC AND KINETIC PROFILES OF YOUR INTERACTION IN A FEW CLICKS
AUTOMATIC RAWDATA PROCESSING
BASELINE CORRECTION &NOISE ELIMINATION
Thermodynamic profileKA = (2.04 ± 0.25)·106 M-1
∆H = (-9.34 ± 0.13) Kcal·mol-1
* KinITC: AN EXCLUSIVE TOOLTO GET KINETIC INFORMATION
THE MODEL BUILDER
An easy to use tool to design your own tailored binding models
GLOBAL FITTING
Perform various experiments and fit globally theresulting isotherms to get the most reliable result
A PERFECT MATCH
Eva Muñoz1, Angel Piñeiro1,2, Juan Sabin1, Javier Rial1, Philippe Dumas3, Eric Ennifar3
2.- GET STRUCTURAL AND MECHANISTIC INFORMATION OF COMPLEX INTERACTING SYSTEMS
3.- QUICK ACCESS TO DATA INTERPRETATION TOOLS
PEAK INTEGRATION
WITH ASSOCIATED ERRORS
Kinetic profilekon = (2.26 ± 0.19)·104 M-1·s-1
koff = (1.11 ± 0.09)·10-2 s-1
RAW DATA
PROCESSSED DATA
ITC ISOTHERM
EQUILIBRATION TIME CURVE (ETC)
M = RECEPTOR IN CELLA = LIGAND IN SYRINGEB = COMPETING LIGAND
COMPETITIVE BINDING OF TWO LIGANDS BINDING TO A THREE SITE RECEPTOR
EQUILIBRATION TIME
LIGAND “A” INTO RECEPTOR LIGAND “B” INTO RECEPTOR LIGAND “A” INTO (RECEPTOR-LIGAND “B”)
Local Minimum
Global Minimum
FRACTION OF OCCUPIED SITES
SITE 1
SITE 2
CONTRIBUTIONS TO THE BINDING ISOTHERM
SITE 1
SITE 3
SITE 2
LIGAND INTO MULTIVALENT POLYMER
LOCAL MINIMA
LIGAND BINDING Nº OF SITES KA (M-1) ∆H (Kcal·mol-1)
Site 1 4-5 (1.93 ± 0.11)·105 -78.14 ± 11.24
Site 2 1-2 (1.68 ± 0.08)·107 -0.14 ± 11.37
Site 3 7-9 (5.30 ± 1.01)·106 -98.44 ± 1.17
SITE 3
References* a) Burnouf D, Ennifar E, Guedich S, Puffer-Enders B, Hoffmann G, Bec G., Disdier F, Baltzinger M, Dumas P (2012) JACS 134, 559-565 b) Dumas P, Ennifar E, Bec G, Piñeiro A, Sabín J, Muñoz E, Rial J, Implementation of KinITC into AFFINImeter, Malvern-MicroCal Application note KinITC: a new method for obtaining joint thermodynamic and kinetic data by isothermal titration calorimetry. (2015).