Fourier transform mass spectrometry to monitor hyaluronan-protein interactions: use of hydrogen/deuterium amide exchange.

Fourier transform mass spectrometry to monitor hyaluronan-protein interactions: use of hydrogen/deuterium amide exchange. Research Abstract Details 

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Fourier transform mass spectrometry to monitor hyaluronan-protein interactions: use of hydrogen/deuterium amide exchange. Abstract Text:

Nicholas T Seyfried,James A Atwood,Austin Yongye,Andrew Almond,Anthony J Day,Ron Orlando,Robert J Woods,

The use of Fourier transform mass spectrometry (FTMS) to monitor noncovalent complex formation in the gas phase under native conditions between the Link module from human tumor necrosis factor stimulated gene-6 (Link_TSG6) and hyaluronan (HA) oligosaccharides is reported. In particular, a titration experiment with increasing concentrations of octasaccharide (HA(8)) to protein produced a noncovalent complex with 1:1 stoichiometry when the oligosaccharide was in molar excess. However, in the presence of a molar excess of tetrasaccharide (HA(4)) nearly all proteins and oligosaccharides were observed in their unbound charge states. These results are consistent with solution-phase properties for this interaction in which HA(8), but not HA(4), supports high affinity Link_TSG6 binding. Hydrogen/deuterium amide exchange mass spectrometry (H/D-EX MS) was also utilized to investigate the level of global deuterium incorporation, over time, for Link_TSG6 in both the absence and presence of HA(8). After dilution into quenching conditions, deuterium incorporation reached limiting asymptotic values of 37 and 26 deuterons for the free and bound protein at 240 and 480 min, respectively, indicating that the oligosaccharide interferes with amide exchange on binding. To detect sequence-specific deuterium incorporation, pepsin digestion of Link_TSG6 in both the absence and presence of HA(8) was performed. A level of deuterium incorporation of 10-30% was observed for peptides analyzed in free Link_TSG6. Interestingly, HA(8) blocked some sites of proteolysis in Link_TSG6 compared to the free protein. Molecular modeling indicated that amino acids proximal to the ligand correlated with regions of the protein that were resistant to enzymatic digestion. Of the peptides that could be analyzed by H/D-EX MS in the presence of the ligand, a 30-60% reduction in deuterium incorporation, relative to the free protein, was observed, even for those sequences not directly involved in HA binding. These results support the utility of FTMS as a method for the characterization of protein-carbohydrate interactions.

Fourier transform mass spectrometry to monitor hyaluronan-protein interactions: use of hydrogen/deuterium amide exchange. Publishing Authors By Initials

NT Seyfried,JA Atwood,A Yongye,A Almond,AJ Day,R Orlando,RJ Woods,

For similar investigative techniques: chemistry, analytical: photometry: spectrophotometry: spectrophotometry, infrared: spectroscopy, fourier transform infrared research abstracts see: investigative techniques: chemistry, analytical: photometry: spectrophotometry: spectrophotometry, infrared: spectroscopy, fourier transform infrared research

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Fourier transform mass spectrometry to monitor hyaluronan-protein interactions: use of hydrogen/deuterium amide exchange. Journal Published:

PUBLICATION TYPE: Research Support, N.I.H., Extr

Journal: Rapid communications in mass spectrometry : RCM

VOLUME: 21

Page Numbers: 121-31

Journal Abbreviation: Rapid Commun. Mass Spectrom.

ISSN: 0951-4198

DAY: 3

MONTH: 12

YEAR: 2007

Fourier transform mass spectrometry to monitor hyaluronan-protein interactions: use of hydrogen/deuterium amide exchange. Information

Number of References:

LANGUAGE: eng

NlmUniqueID: 8802365

Fourier transform mass spectrometry to monitor hyaluronan-protein interactions: use of hydrogen/deuterium amide exchange. Keywords Mesh Terms:

KEYWORDS: Spectroscopy, Fourier Transform Infrared

MESH TERMS: methods

Chemical & Substance for Abstract: Fourier transform mass spectrometry to monitor hyaluronan-protein interactions: use of hydrogen/deuterium amide exchange. Information

Substance Name: Hyaluronic Acid

Registry Number: 9004-61-9

Grant and Affiliation Information for Fourier transform mass spectrometry to monitor hyaluronan-protein interactions: use of hydrogen/deuterium amide exchange.

AFFILIATION: Complex Carbohydrate Research Center, University of Georgia, 315 Riverbend Road, Athens, GA 30602, USA.

Country: England

AGENCY: United States NCRR

GRANT: RR 0537

ACRONYM: RR

MEDLINETA: Rapid Commun Mass Spectrom

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