Isothermal titration calorimetry

In summary, the conversation discusses the use of isothermal titration calorimetry to calculate \DeltaH^{o}_{bind} for a protein-ligand binding reaction at 25°C. The speaker also mentions performing a separate assay at different temperatures to measure equilibrium ligand binding. The question posed is how to find \DeltaS^{0}_{bind} at 37C. The speaker suggests using the van't hoff equation to find \DeltaH at 37C since the different Kds and temperatures are already known.
  • #1
Quickdry135
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Homework Statement



Using isothermal titration calorimetry, you calculate [tex]\Delta[/tex]H[tex]^{o}_{bind}[/tex] (= -5000 cal/mol) for a protein-ligand binding reaction at 25°C. You then perform a separate assay in which you measure equilibrium ligand binding at two different temperatures:

L[tex]_{0}[/tex] (nM) Ceq (nM) at 25°C Ceq (nM) at 37°C
0.01 :: 0.007 :: 0.006
0.03 :: 0.021 :: 0.017
0.1 ;: 0.070 :: 0.058
0.3 :: 0.197 ;: 0.166
1 :: 0.537 :: 0.439
3 ;: 0.830 :: 0.778
10 :: 0.943 :: 0.930
30 :: 1.002 :: 0.964
100 :: 0.981 :: 1.009

What is [tex]\Delta[/tex]S[tex]^{0}_{bind}[/tex] at 37C

Homework Equations


[tex]\Delta[/tex]G[tex]^{0}[/tex]=RTlnK[tex]_{D}[/tex]

[tex]\Delta[/tex]G=[tex]\Delta[/tex]G[tex]^{0}[/tex] + RTln[L][tex]_{eq}[/tex]/[P][tex]_{eq}[/tex][L][tex]_{eq}[/tex]

[tex]\Delta[/tex]G=[tex]\Delta[/tex]H-T[tex]\Delta[/tex]S


The Attempt at a Solution



I can find K[tex]_{d}[/tex] graphically and therefore find [tex]\Delta[/tex]G[tex]^{0}[/tex] to be -12300.1 cal. At the same temperature, [tex]\Delta[/tex]G[tex]^{0}[/tex]=[tex]\Delta[/tex]G at 25C, so i can find \DeltaS at 25C. But I don't know how this helps me find [tex]\Delta[/tex]S[tex]^{0}_{bind}[/tex] at 37C or if this helps me at all.

thanks for any help or direction
 
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  • #2
never mind, I think i figured it out using the van't hoff equation to find delta H at 37C since I can find the different Kds and am given the different temperatures.
 
  • #3
you can give me


I would respond by saying that the data provided suggests that the binding reaction is exothermic (\DeltaH^{o}_{bind} = -5000 cal/mol) and that the change in enthalpy and entropy at 25°C can be calculated using the equations \DeltaG^{0}=RTlnK_{D} and \DeltaG=\DeltaH-T\DeltaS. However, in order to calculate \DeltaS^{0}_{bind} at 37°C, we would need to also know the change in enthalpy at that temperature. This can be calculated using the relationship \DeltaH^{o}_{bind} = \DeltaH^{o}_{bind} + \DeltaC_{p}\DeltaT, where \DeltaC_{p} is the change in heat capacity and \DeltaT is the change in temperature (37°C - 25°C). Once we have the value for \DeltaH^{o}_{bind} at 37°C, we can use the equation \DeltaG=\DeltaH-T\DeltaS to calculate \DeltaS^{0}_{bind} at that temperature. It would also be helpful to have more data points at different temperatures in order to accurately calculate the change in enthalpy and entropy over a range of temperatures.
 

What is isothermal titration calorimetry?

Isothermal titration calorimetry (ITC) is a technique used to measure the heat released or absorbed during a chemical reaction or binding event. It is based on the principle that any change in enthalpy (heat content) produces a corresponding change in temperature.

How does isothermal titration calorimetry work?

In ITC, a known amount of one reactant is titrated into a solution containing the other reactant. As the reaction occurs, the heat released or absorbed is measured by a sensitive calorimeter. This heat is then plotted against the molar ratio of the two reactants, allowing for the determination of the binding affinity and thermodynamic parameters of the reaction.

What types of interactions can be studied using isothermal titration calorimetry?

ITC can be used to study a variety of interactions, including protein-ligand binding, receptor-ligand binding, enzyme-substrate interactions, and antibody-antigen binding. It can also be used to study the binding of small molecules to nucleic acids or lipids.

What are the advantages of using isothermal titration calorimetry?

ITC is a label-free technique that does not require any modification of the molecules being studied. It also provides quantitative information on the thermodynamics of the interaction, including binding affinity, enthalpy, and entropy. Additionally, ITC is a fast and sensitive technique, requiring only small amounts of sample.

What are the limitations of isothermal titration calorimetry?

ITC is limited by the size of the molecules being studied, as very large or small molecules may not produce a measurable heat signal. It also requires the molecules to be soluble and stable in solution. Additionally, ITC can only provide information on the overall binding reaction and cannot differentiate between different binding sites or binding modes.

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