MS, with or without pre-analysis peptide fractionation, can be used to decipher the residues on proteins where oxidative modifications caused by peroxynitrite, singlet oxygen or electrophilic lipids have occurred. Peroxynitrite nitrates tyrosine and tryptophan residues on the surface of actin. Singlet oxygen, formed by the interaction of UVA light with tryptophan, can oxidize neighbouring cysteine, histidine, methionine, tyrosine and tryptophan residues. Dose–response inactivation by 4HNE (4-hydroxynonenal) of hBAT (human bile acid CoA:amino acid N-acyltransferase) and CKBB (cytosolic brain isoform of creatine kinase) is associated with site-specific modifications. FT-ICR (Fourier-transform ion cyclotron resonance)–MS using nanoLC (nano-liquid chromatography)–ESI (electrospray ionization)–MS or direct-infusion ESI–MS with gas-phase fractionation identified 14 4HNE adducts on hBAT and 17 on CKBB respectively. At 4HNE concentrations in the physiological range, one member of the catalytic triad of hBAT (His362) was modified; for CKBB, although all four residues in the active site that were modifiable by 4HNE were ultimately modified, only one, Cys283, occurred at physiological concentrations of 4HNE. These results suggest that future in vivo studies should carefully assess the critical sites that are modified rather than using antibodies that do not distinguish between different modified sites.
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October 2008
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Conference Article|
September 19 2008
High-resolution mass spectrometry analysis of protein oxidations and resultant loss of function
Stephen Barnes;
Stephen Barnes
1
*Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
†Department of Pharmacology and Toxicology, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
‡Department of Vision Sciences, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
§UAB Center for Nutrient-Gene Interaction, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
∥UAB Laboratory for Biomedical FT-ICR Mass Spectrometry, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
¶Purdue University–University of Alabama at Birmingham Botanicals Center for Age-Related Disease, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
1To whom correspondence should be addressed (email [email protected]).
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Erin M. Shonsey;
Erin M. Shonsey
†Department of Pharmacology and Toxicology, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
∥UAB Laboratory for Biomedical FT-ICR Mass Spectrometry, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
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Shannon M. Eliuk;
Shannon M. Eliuk
†Department of Pharmacology and Toxicology, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
¶Purdue University–University of Alabama at Birmingham Botanicals Center for Age-Related Disease, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
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David Stella;
David Stella
*Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
¶Purdue University–University of Alabama at Birmingham Botanicals Center for Age-Related Disease, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
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Kerri Barrett;
Kerri Barrett
2
¶Purdue University–University of Alabama at Birmingham Botanicals Center for Age-Related Disease, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
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Om P. Srivastava;
Om P. Srivastava
‡Department of Vision Sciences, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
¶Purdue University–University of Alabama at Birmingham Botanicals Center for Age-Related Disease, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
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Helen Kim;
Helen Kim
*Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
†Department of Pharmacology and Toxicology, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
§UAB Center for Nutrient-Gene Interaction, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
¶Purdue University–University of Alabama at Birmingham Botanicals Center for Age-Related Disease, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
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Matthew B. Renfrow
Matthew B. Renfrow
*Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
∥UAB Laboratory for Biomedical FT-ICR Mass Spectrometry, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
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Publisher: Portland Press Ltd
Received:
May 29 2008
Online ISSN: 1470-8752
Print ISSN: 0300-5127
© The Authors Journal compilation © 2008 Biochemical Society
2008
Biochem Soc Trans (2008) 36 (5): 1037–1044.
Article history
Received:
May 29 2008
Citation
Stephen Barnes, Erin M. Shonsey, Shannon M. Eliuk, David Stella, Kerri Barrett, Om P. Srivastava, Helen Kim, Matthew B. Renfrow; High-resolution mass spectrometry analysis of protein oxidations and resultant loss of function. Biochem Soc Trans 1 October 2008; 36 (5): 1037–1044. doi: https://doi.org/10.1042/BST0361037
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