Low-affinity binding determined by titration calorimetry using a high- affinity coupling ligand: A thermodynamic study of ligand binding to protein tyrosine phosphatase 1B

Yan Ling Zhang, Zhong-Yin Zhang

Research output: Contribution to journalArticle

121 Citations (Scopus)

Abstract

A competition-based method is used for the determination of the thermodynamic parameters for a low-affinity ligand binding reaction by isothermal titration calorimetry. This method is based on the coupling of a high-affinity ligand to the binding of the low-affinity ligand. Results are presented for the binding of a nonhydrolyzable phosphotyrosine analog phosphonodifluoromethyl phenylalanine (F2Pmp)-containing peptide (Ac-Asp- Ala-Asp-Glu-F2Pmp-Leu-NH2), arsenate, and inorganic phosphate to the intracellular human protein tyrosine phosphatase 1B (PTP1B). The binding constants are 3.3 x 106, 4.3 x 103, and 48 M-1 for the F2Pmp-containing peptide, arsenate, and inorganic phosphate, respectively. The binding of arsenate and inorganic phosphate to PTP1B is enthalpy driven. This is in contrast to the binding of the F2Pmp-containing peptide which is mainly driven by entropy. The calorimetrically determined binding constants are in agreement with the K(i) values determined by enzyme inhibition studies. This demonstrates that isothermal titration calorimetry can be used to quantitatively determine the thermodynamic parameters for the interactions between proteins and low-affinity ligands if a proper coupling ligand can be identified.

Original languageEnglish (US)
Pages (from-to)139-148
Number of pages10
JournalAnalytical Biochemistry
Volume261
Issue number2
DOIs
StatePublished - Aug 1 1998
Externally publishedYes

Fingerprint

Non-Receptor Type 1 Protein Tyrosine Phosphatase
Calorimetry
Titration
Thermodynamics
Carrier Proteins
Ligands
Phosphates
Peptides
Enzyme inhibition
Phosphotyrosine
Entropy
Phenylalanine
Enthalpy
Enzymes
arsenic acid
Proteins

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

@article{3a5185a94c3a4660a6534e53ab6828ba,
title = "Low-affinity binding determined by titration calorimetry using a high- affinity coupling ligand: A thermodynamic study of ligand binding to protein tyrosine phosphatase 1B",
abstract = "A competition-based method is used for the determination of the thermodynamic parameters for a low-affinity ligand binding reaction by isothermal titration calorimetry. This method is based on the coupling of a high-affinity ligand to the binding of the low-affinity ligand. Results are presented for the binding of a nonhydrolyzable phosphotyrosine analog phosphonodifluoromethyl phenylalanine (F2Pmp)-containing peptide (Ac-Asp- Ala-Asp-Glu-F2Pmp-Leu-NH2), arsenate, and inorganic phosphate to the intracellular human protein tyrosine phosphatase 1B (PTP1B). The binding constants are 3.3 x 106, 4.3 x 103, and 48 M-1 for the F2Pmp-containing peptide, arsenate, and inorganic phosphate, respectively. The binding of arsenate and inorganic phosphate to PTP1B is enthalpy driven. This is in contrast to the binding of the F2Pmp-containing peptide which is mainly driven by entropy. The calorimetrically determined binding constants are in agreement with the K(i) values determined by enzyme inhibition studies. This demonstrates that isothermal titration calorimetry can be used to quantitatively determine the thermodynamic parameters for the interactions between proteins and low-affinity ligands if a proper coupling ligand can be identified.",
author = "Zhang, {Yan Ling} and Zhong-Yin Zhang",
year = "1998",
month = "8",
day = "1",
doi = "10.1006/abio.1998.2738",
language = "English (US)",
volume = "261",
pages = "139--148",
journal = "Analytical Biochemistry",
issn = "0003-2697",
publisher = "Academic Press Inc.",
number = "2",

}

TY - JOUR

T1 - Low-affinity binding determined by titration calorimetry using a high- affinity coupling ligand

T2 - A thermodynamic study of ligand binding to protein tyrosine phosphatase 1B

AU - Zhang, Yan Ling

AU - Zhang, Zhong-Yin

PY - 1998/8/1

Y1 - 1998/8/1

N2 - A competition-based method is used for the determination of the thermodynamic parameters for a low-affinity ligand binding reaction by isothermal titration calorimetry. This method is based on the coupling of a high-affinity ligand to the binding of the low-affinity ligand. Results are presented for the binding of a nonhydrolyzable phosphotyrosine analog phosphonodifluoromethyl phenylalanine (F2Pmp)-containing peptide (Ac-Asp- Ala-Asp-Glu-F2Pmp-Leu-NH2), arsenate, and inorganic phosphate to the intracellular human protein tyrosine phosphatase 1B (PTP1B). The binding constants are 3.3 x 106, 4.3 x 103, and 48 M-1 for the F2Pmp-containing peptide, arsenate, and inorganic phosphate, respectively. The binding of arsenate and inorganic phosphate to PTP1B is enthalpy driven. This is in contrast to the binding of the F2Pmp-containing peptide which is mainly driven by entropy. The calorimetrically determined binding constants are in agreement with the K(i) values determined by enzyme inhibition studies. This demonstrates that isothermal titration calorimetry can be used to quantitatively determine the thermodynamic parameters for the interactions between proteins and low-affinity ligands if a proper coupling ligand can be identified.

AB - A competition-based method is used for the determination of the thermodynamic parameters for a low-affinity ligand binding reaction by isothermal titration calorimetry. This method is based on the coupling of a high-affinity ligand to the binding of the low-affinity ligand. Results are presented for the binding of a nonhydrolyzable phosphotyrosine analog phosphonodifluoromethyl phenylalanine (F2Pmp)-containing peptide (Ac-Asp- Ala-Asp-Glu-F2Pmp-Leu-NH2), arsenate, and inorganic phosphate to the intracellular human protein tyrosine phosphatase 1B (PTP1B). The binding constants are 3.3 x 106, 4.3 x 103, and 48 M-1 for the F2Pmp-containing peptide, arsenate, and inorganic phosphate, respectively. The binding of arsenate and inorganic phosphate to PTP1B is enthalpy driven. This is in contrast to the binding of the F2Pmp-containing peptide which is mainly driven by entropy. The calorimetrically determined binding constants are in agreement with the K(i) values determined by enzyme inhibition studies. This demonstrates that isothermal titration calorimetry can be used to quantitatively determine the thermodynamic parameters for the interactions between proteins and low-affinity ligands if a proper coupling ligand can be identified.

UR - http://www.scopus.com/inward/record.url?scp=0032144872&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0032144872&partnerID=8YFLogxK

U2 - 10.1006/abio.1998.2738

DO - 10.1006/abio.1998.2738

M3 - Article

C2 - 9716416

AN - SCOPUS:0032144872

VL - 261

SP - 139

EP - 148

JO - Analytical Biochemistry

JF - Analytical Biochemistry

SN - 0003-2697

IS - 2

ER -