Partial colocalization of oxidized, N-formylkynurenine-containing proteins in mitochondria and Golgi of keratinocytes. Photochem Photobiol 2010;86(4):752-6
Date
04/23/2010Pubmed ID
20408979Pubmed Central ID
PMC3615536DOI
10.1111/j.1751-1097.2010.00718.xScopus ID
2-s2.0-77954388280 (requires institutional sign-in at Scopus site) 5 CitationsAbstract
Proteins are the dominant cellular target for oxidative reactions because they comprise the majority of macromolecules. Posttranslational oxidative protein modifications include fragmentation, aggregation and alteration of specific amino acid residues. The amino acids and amino acid residues most susceptible to oxidative modification are those containing sulfur and those with aromatic rings. Tryptophan reacts with radicals, ozone and singlet oxygen to form the end product N-formylkynurenine (NFK). We recently described a novel anti-NFK antiserum and validated its use in immunological assays for the specific detection of NFK in isolated proteins and protein mixtures. Here we photo-oxidize rose bengal-containing HaCaT keratinocyte cells and examine the results using fluorescent confocal microscopy and staining with anti-NFK antiserum and markers for both Golgi and mitochondria. We show that photosensitization mediates the accumulation of NFK and that NFK can be detected in photosensitized cells with only slightly decreased viability. Additionally, we detect NFK-modified proteins in both Golgi and mitochondria of photosensitized cells. These experiments demonstrate that we have developed a tool for the specific detection of oxidized tryptophan residues in cells and suggest that this tool could be useful in tracking the fate of these oxidized proteins.
Author List
Ehrenshaft M, Bonini MG, Feng L, Chignell CF, Mason RPMESH terms used to index this publication - Major topics in bold
Cells, CulturedGolgi Apparatus
Humans
Keratinocytes
Kynurenine
Light
Mitochondria
Oxidation-Reduction
Proteins
Singlet Oxygen
