Electrospray mass spectrometric investigation of the binding of cis-parinaric acid to bovine beta-lactoglobulin and study of the ligand-binding site of the protein using limited proteolysis

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Abstract

The binding property of parinaric acid, a polyunsaturated fatty acid, to bovine β-lactoglobulin, has been studied by electrospray ionization mass spectrometry. Stable complexation was observed under acidic conditions in a molar ratio of 1:1. Competitive complexation experiments were performed using saturated and unsaturated fatty acid standards with different chain lengths and number of double bonds to study the specificity of the interaction. It can be concluded that formation of the parinaric acid-lactoglobulin complex is preferred even if the molar concentration of the other fatty acids is ten times higher. In cases of specific complex formation the protein must have an active site that is a good acceptor for the ligand molecule. Limited trypsinolysis was performed on the lactoglobulin molecule to identify which part is responsible for the complexation. An intermediate tryptic fragment with molecular mass of 5200 Da was found to have the same ability to bind parinaric acid as the intact protein. This disulfide-bonded residue, [41-70]S-S[149-162], might thus be involved in the specific complexation of parinaric acid to β-lactoglobulin. This conclusion is consistent with previous information on this binding site.

Original languageEnglish
Pages (from-to)2464-2470
Number of pages7
JournalRapid Communications in Mass Spectrometry
Volume17
Issue number22
Publication statusPublished - 2003

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Proteolysis
Lactoglobulins
Complexation
Binding Sites
Ligands
Unsaturated Fatty Acids
Proteins
Fatty Acids
Electrospray ionization
Molecules
Molecular mass
Chain length
Disulfides
Mass spectrometry
parinaric acid
Experiments

ASJC Scopus subject areas

  • Analytical Chemistry
  • Spectroscopy

Cite this

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title = "Electrospray mass spectrometric investigation of the binding of cis-parinaric acid to bovine beta-lactoglobulin and study of the ligand-binding site of the protein using limited proteolysis",
abstract = "The binding property of parinaric acid, a polyunsaturated fatty acid, to bovine β-lactoglobulin, has been studied by electrospray ionization mass spectrometry. Stable complexation was observed under acidic conditions in a molar ratio of 1:1. Competitive complexation experiments were performed using saturated and unsaturated fatty acid standards with different chain lengths and number of double bonds to study the specificity of the interaction. It can be concluded that formation of the parinaric acid-lactoglobulin complex is preferred even if the molar concentration of the other fatty acids is ten times higher. In cases of specific complex formation the protein must have an active site that is a good acceptor for the ligand molecule. Limited trypsinolysis was performed on the lactoglobulin molecule to identify which part is responsible for the complexation. An intermediate tryptic fragment with molecular mass of 5200 Da was found to have the same ability to bind parinaric acid as the intact protein. This disulfide-bonded residue, [41-70]S-S[149-162], might thus be involved in the specific complexation of parinaric acid to β-lactoglobulin. This conclusion is consistent with previous information on this binding site.",
author = "T. Imre and F. Zsila and P. Szab{\'o}",
year = "2003",
language = "English",
volume = "17",
pages = "2464--2470",
journal = "Rapid Communications in Mass Spectrometry",
issn = "0951-4198",
publisher = "John Wiley and Sons Ltd",
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}

TY - JOUR

T1 - Electrospray mass spectrometric investigation of the binding of cis-parinaric acid to bovine beta-lactoglobulin and study of the ligand-binding site of the protein using limited proteolysis

AU - Imre, T.

AU - Zsila, F.

AU - Szabó, P.

PY - 2003

Y1 - 2003

N2 - The binding property of parinaric acid, a polyunsaturated fatty acid, to bovine β-lactoglobulin, has been studied by electrospray ionization mass spectrometry. Stable complexation was observed under acidic conditions in a molar ratio of 1:1. Competitive complexation experiments were performed using saturated and unsaturated fatty acid standards with different chain lengths and number of double bonds to study the specificity of the interaction. It can be concluded that formation of the parinaric acid-lactoglobulin complex is preferred even if the molar concentration of the other fatty acids is ten times higher. In cases of specific complex formation the protein must have an active site that is a good acceptor for the ligand molecule. Limited trypsinolysis was performed on the lactoglobulin molecule to identify which part is responsible for the complexation. An intermediate tryptic fragment with molecular mass of 5200 Da was found to have the same ability to bind parinaric acid as the intact protein. This disulfide-bonded residue, [41-70]S-S[149-162], might thus be involved in the specific complexation of parinaric acid to β-lactoglobulin. This conclusion is consistent with previous information on this binding site.

AB - The binding property of parinaric acid, a polyunsaturated fatty acid, to bovine β-lactoglobulin, has been studied by electrospray ionization mass spectrometry. Stable complexation was observed under acidic conditions in a molar ratio of 1:1. Competitive complexation experiments were performed using saturated and unsaturated fatty acid standards with different chain lengths and number of double bonds to study the specificity of the interaction. It can be concluded that formation of the parinaric acid-lactoglobulin complex is preferred even if the molar concentration of the other fatty acids is ten times higher. In cases of specific complex formation the protein must have an active site that is a good acceptor for the ligand molecule. Limited trypsinolysis was performed on the lactoglobulin molecule to identify which part is responsible for the complexation. An intermediate tryptic fragment with molecular mass of 5200 Da was found to have the same ability to bind parinaric acid as the intact protein. This disulfide-bonded residue, [41-70]S-S[149-162], might thus be involved in the specific complexation of parinaric acid to β-lactoglobulin. This conclusion is consistent with previous information on this binding site.

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M3 - Article

VL - 17

SP - 2464

EP - 2470

JO - Rapid Communications in Mass Spectrometry

JF - Rapid Communications in Mass Spectrometry

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