Proton Nuclear Magnetic Resonance Spectroscopy and Ligand Binding Dynamics of the Escherichia coli l-Arabinose Binding Protein

Abbot Clark, Robert W. Hogg, Thomas A. Gerken

Research output: Contribution to journalArticle

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Abstract

The l-arabinose binding protein (ABP) from Escherichia coli was studied by proton nuclear magnetic resonance spectroscopy (1H NMR). Distinct spectral changes occur when ABP binds its natural ligand, l-arabinose, which involve resonances in the aromatic ring current shifted methyl, bulk methyl, methylene, aromatic, and amide proton regions of the spectra. Several amide resonances can be “protected” from deuterium exchange if l-arabinose is bound to ABP prior to deuterium oxide dialysis. On the basis of the pH dependence of their chemical shifts, two low-field resonances have been tentatively assigned to C2 protons of two of the three histidines present in ABP. These histidyl residues have pK values of 8.0 and 8.6 which support their involvement in ionic interactions observed earlier in the crystallographic analysis. One histidyl residue shows a small chemical shift change upon the addition of arabinose. When ABP binds d-galactose, changes in the spectra occur which are different than those observed when l-arabinose is bound. Binding of l-arabinose and d-galactose to the binding protein (ABP) was considered by equilibrium binding and fluorescence emission spectroscopy. ABP binds l-arabinose and d-galactose with high affinities (Kd's at 6 °C of 1.3 × 10−7 and 1.9 × 10−7 M, respectively), and both enthalpy and entropy contribute to the ABP-ligand association. When excited at 285 nm, ABP has a fluorescence emission maximum of 340 nm which is quenched and blue shifted (to 337 nm) upon binding l-arabinose. ABP binding d-galactose produced a similar emission shift but no fluorescence quenching.

Original languageEnglish
Pages (from-to)2227-2233
Number of pages7
JournalBiochemistry
Volume21
Issue number9
DOIs
StatePublished - 1 Apr 1982

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Arabinose
Escherichia coli
Nuclear magnetic resonance spectroscopy
Carrier Proteins
Magnetic Resonance Spectroscopy
Nuclear magnetic resonance
Ligands
Galactose
Proton Magnetic Resonance Spectroscopy
Chemical shift
Amides
Protons
Fluorescence
Deuterium Oxide
Dialysis
Deuterium
Fluorescence Spectrometry
Fluorescence spectroscopy
Emission spectroscopy
Entropy

Cite this

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title = "Proton Nuclear Magnetic Resonance Spectroscopy and Ligand Binding Dynamics of the Escherichia coli l-Arabinose Binding Protein",
abstract = "The l-arabinose binding protein (ABP) from Escherichia coli was studied by proton nuclear magnetic resonance spectroscopy (1H NMR). Distinct spectral changes occur when ABP binds its natural ligand, l-arabinose, which involve resonances in the aromatic ring current shifted methyl, bulk methyl, methylene, aromatic, and amide proton regions of the spectra. Several amide resonances can be “protected” from deuterium exchange if l-arabinose is bound to ABP prior to deuterium oxide dialysis. On the basis of the pH dependence of their chemical shifts, two low-field resonances have been tentatively assigned to C2 protons of two of the three histidines present in ABP. These histidyl residues have pK values of 8.0 and 8.6 which support their involvement in ionic interactions observed earlier in the crystallographic analysis. One histidyl residue shows a small chemical shift change upon the addition of arabinose. When ABP binds d-galactose, changes in the spectra occur which are different than those observed when l-arabinose is bound. Binding of l-arabinose and d-galactose to the binding protein (ABP) was considered by equilibrium binding and fluorescence emission spectroscopy. ABP binds l-arabinose and d-galactose with high affinities (Kd's at 6 °C of 1.3 × 10−7 and 1.9 × 10−7 M, respectively), and both enthalpy and entropy contribute to the ABP-ligand association. When excited at 285 nm, ABP has a fluorescence emission maximum of 340 nm which is quenched and blue shifted (to 337 nm) upon binding l-arabinose. ABP binding d-galactose produced a similar emission shift but no fluorescence quenching.",
author = "Abbot Clark and Hogg, {Robert W.} and Gerken, {Thomas A.}",
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Proton Nuclear Magnetic Resonance Spectroscopy and Ligand Binding Dynamics of the Escherichia coli l-Arabinose Binding Protein. / Clark, Abbot; Hogg, Robert W.; Gerken, Thomas A.

In: Biochemistry, Vol. 21, No. 9, 01.04.1982, p. 2227-2233.

Research output: Contribution to journalArticle

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AU - Clark, Abbot

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