A Linear Correlation between the Energetics of Allosteric Communication and Protein Flexibility in the Escherichia coli Cyclic AMP Receptor Protein Revealed by Mutation-Induced Changes in Compressibility and Amide Hydrogen-Deuterium Exchange

Kunihiko Gekko, Norihiro Obu, Jianquan Li, James Lee

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Abstract

Amino acid substitutions at distant sites in the Escherichia coli cyclic AMP receptor protein (CRP) have been shown to affect both the nature and magnitude of the energetics of cooperativity of cAMP binding, ranging from negative to positive. In addition, the binding to DNA is concomitantly affected. To correlate the effects of amino acid substitutions on the functional energetics and global structural properties in CRP, the partial specific volume (v̄°), the coefficient of adiabatic compressibility (β̄ s°), and the rate of amide proton exchange were determined for the wild-type and eight mutant CRPs (K52N, D53H, S62F, T127L, G141Q, L148R, H159L, and K52N/H159L) by using sound velocity, density measurements, and hydrogen-deuterium exchange as monitored by Fourier transform infrared spectroscopy at 25 °C. These mutations induced large changes in v̄° (0.747-0.756 mL/g) and β̄s° (6.89-9.68 Mbar-1) compared to the corresponding values for wild-type CRP (v̄° = 0.750 mL/g and β̄s° = 7.98 Mbar -1). These changes in global structural properties correlated with the rate of amide proton exchange. A linear correlation was established between β̄s° and the energetics of cooperativity of binding of cAMP to the high-affinity sites, regardless of the nature of cooperativity, be it negative or positive. This linear correlation indicates that the nature and magnitude of cooperativity are a continuum. A similar linear correlation was established between compressibility and DNA binding affinity. In addition, linear correlations were also found among the dynamics of CRP and functional energetics. Double mutation (K52N/H159L) at positions 52 and 159, whose α-carbons are separated by 34.6 Å, showed nonadditive effects on v̄° and β̄s°. These results demonstrate that a small alteration in the local structure due to amino acid substitution is dramatically magnified in the overall protein dynamics which plays an important role in modulating the allosteric behavior of CRP.

Original languageEnglish (US)
Pages (from-to)3844-3852
Number of pages9
JournalBiochemistry
Volume43
Issue number13
DOIs
StatePublished - Apr 6 2004

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Cyclic AMP Receptor Protein
Deuterium
Escherichia coli Proteins
Compressibility
Amides
Escherichia coli
Hydrogen
Ion exchange
Amino Acid Substitution
Mutation
Communication
Substitution reactions
Proteins
Amino Acids
Deuterium Exchange Measurement
Protons
Structural properties
DNA
Acoustic wave velocity
Fourier Transform Infrared Spectroscopy

ASJC Scopus subject areas

  • Biochemistry

Cite this

@article{99543fed413145d2a0882b3f8bf3a77b,
title = "A Linear Correlation between the Energetics of Allosteric Communication and Protein Flexibility in the Escherichia coli Cyclic AMP Receptor Protein Revealed by Mutation-Induced Changes in Compressibility and Amide Hydrogen-Deuterium Exchange",
abstract = "Amino acid substitutions at distant sites in the Escherichia coli cyclic AMP receptor protein (CRP) have been shown to affect both the nature and magnitude of the energetics of cooperativity of cAMP binding, ranging from negative to positive. In addition, the binding to DNA is concomitantly affected. To correlate the effects of amino acid substitutions on the functional energetics and global structural properties in CRP, the partial specific volume (v̄°), the coefficient of adiabatic compressibility (β̄ s°), and the rate of amide proton exchange were determined for the wild-type and eight mutant CRPs (K52N, D53H, S62F, T127L, G141Q, L148R, H159L, and K52N/H159L) by using sound velocity, density measurements, and hydrogen-deuterium exchange as monitored by Fourier transform infrared spectroscopy at 25 °C. These mutations induced large changes in v̄° (0.747-0.756 mL/g) and β̄s° (6.89-9.68 Mbar-1) compared to the corresponding values for wild-type CRP (v̄° = 0.750 mL/g and β̄s° = 7.98 Mbar -1). These changes in global structural properties correlated with the rate of amide proton exchange. A linear correlation was established between β̄s° and the energetics of cooperativity of binding of cAMP to the high-affinity sites, regardless of the nature of cooperativity, be it negative or positive. This linear correlation indicates that the nature and magnitude of cooperativity are a continuum. A similar linear correlation was established between compressibility and DNA binding affinity. In addition, linear correlations were also found among the dynamics of CRP and functional energetics. Double mutation (K52N/H159L) at positions 52 and 159, whose α-carbons are separated by 34.6 {\AA}, showed nonadditive effects on v̄° and β̄s°. These results demonstrate that a small alteration in the local structure due to amino acid substitution is dramatically magnified in the overall protein dynamics which plays an important role in modulating the allosteric behavior of CRP.",
author = "Kunihiko Gekko and Norihiro Obu and Jianquan Li and James Lee",
year = "2004",
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doi = "10.1021/bi036271e",
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T1 - A Linear Correlation between the Energetics of Allosteric Communication and Protein Flexibility in the Escherichia coli Cyclic AMP Receptor Protein Revealed by Mutation-Induced Changes in Compressibility and Amide Hydrogen-Deuterium Exchange

AU - Gekko, Kunihiko

AU - Obu, Norihiro

AU - Li, Jianquan

AU - Lee, James

PY - 2004/4/6

Y1 - 2004/4/6

N2 - Amino acid substitutions at distant sites in the Escherichia coli cyclic AMP receptor protein (CRP) have been shown to affect both the nature and magnitude of the energetics of cooperativity of cAMP binding, ranging from negative to positive. In addition, the binding to DNA is concomitantly affected. To correlate the effects of amino acid substitutions on the functional energetics and global structural properties in CRP, the partial specific volume (v̄°), the coefficient of adiabatic compressibility (β̄ s°), and the rate of amide proton exchange were determined for the wild-type and eight mutant CRPs (K52N, D53H, S62F, T127L, G141Q, L148R, H159L, and K52N/H159L) by using sound velocity, density measurements, and hydrogen-deuterium exchange as monitored by Fourier transform infrared spectroscopy at 25 °C. These mutations induced large changes in v̄° (0.747-0.756 mL/g) and β̄s° (6.89-9.68 Mbar-1) compared to the corresponding values for wild-type CRP (v̄° = 0.750 mL/g and β̄s° = 7.98 Mbar -1). These changes in global structural properties correlated with the rate of amide proton exchange. A linear correlation was established between β̄s° and the energetics of cooperativity of binding of cAMP to the high-affinity sites, regardless of the nature of cooperativity, be it negative or positive. This linear correlation indicates that the nature and magnitude of cooperativity are a continuum. A similar linear correlation was established between compressibility and DNA binding affinity. In addition, linear correlations were also found among the dynamics of CRP and functional energetics. Double mutation (K52N/H159L) at positions 52 and 159, whose α-carbons are separated by 34.6 Å, showed nonadditive effects on v̄° and β̄s°. These results demonstrate that a small alteration in the local structure due to amino acid substitution is dramatically magnified in the overall protein dynamics which plays an important role in modulating the allosteric behavior of CRP.

AB - Amino acid substitutions at distant sites in the Escherichia coli cyclic AMP receptor protein (CRP) have been shown to affect both the nature and magnitude of the energetics of cooperativity of cAMP binding, ranging from negative to positive. In addition, the binding to DNA is concomitantly affected. To correlate the effects of amino acid substitutions on the functional energetics and global structural properties in CRP, the partial specific volume (v̄°), the coefficient of adiabatic compressibility (β̄ s°), and the rate of amide proton exchange were determined for the wild-type and eight mutant CRPs (K52N, D53H, S62F, T127L, G141Q, L148R, H159L, and K52N/H159L) by using sound velocity, density measurements, and hydrogen-deuterium exchange as monitored by Fourier transform infrared spectroscopy at 25 °C. These mutations induced large changes in v̄° (0.747-0.756 mL/g) and β̄s° (6.89-9.68 Mbar-1) compared to the corresponding values for wild-type CRP (v̄° = 0.750 mL/g and β̄s° = 7.98 Mbar -1). These changes in global structural properties correlated with the rate of amide proton exchange. A linear correlation was established between β̄s° and the energetics of cooperativity of binding of cAMP to the high-affinity sites, regardless of the nature of cooperativity, be it negative or positive. This linear correlation indicates that the nature and magnitude of cooperativity are a continuum. A similar linear correlation was established between compressibility and DNA binding affinity. In addition, linear correlations were also found among the dynamics of CRP and functional energetics. Double mutation (K52N/H159L) at positions 52 and 159, whose α-carbons are separated by 34.6 Å, showed nonadditive effects on v̄° and β̄s°. These results demonstrate that a small alteration in the local structure due to amino acid substitution is dramatically magnified in the overall protein dynamics which plays an important role in modulating the allosteric behavior of CRP.

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