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Detection and localization of oxidized proteins in muscle cells by fluorescence microscopy
- Source :
- Journal of Agricultural and Food Chemistry, Journal of Agricultural and Food Chemistry, American Chemical Society, 2007, 55 (23), pp.9554-9558. ⟨10.1021/jf0717586⟩
- Publication Year :
- 2007
-
Abstract
- International audience; In meat, no detailed studies on the intracellular distribution of oxidized proteins during oxidative stress have been performed, to our knowledge. Therefore, we used fluorescence microscopy to detect and locate protein carbonyls, oxidation products of basic amino acids, generated in bovine M. Rectus abdominis during either exposition to a chemical free radical generating system, or refrigerated storage, or cooking. The technique consisted of an immunohistochemical detection of carbonyls by reaction with the specific probe DNPH (2,4-dinitrophenylhydrazine) followed by the sequential addition of a first antibody against DNPH-carbonylated proteins and a CY3-labeled secondary antibody. The fluorescence of the CY3 probe increased regularly with level of free radical generating system and storage time. Moreover, an important heterogeneity of carbonyl distribution was observed, with a higher oxidation level at the periphery than inside the muscle cells. Cooking induced fluorescence increase only at the periphery of cells. Specific coloration of collagen by Sirius red showed that collagen was not involved in fluorescence. We can deduce that accumulation of oxidized proteins observed in the cell periphery was linked to membrane protein oxidation and not to connective tissue oxidation. Biochemical assays were performed in parallel on membrane and myofibrillar proteins to provide complementary quantitative data on level of oxidized proteins.
- Subjects :
- Hot Temperature
Meat
Free Radicals
Muscle Proteins
Protein oxidation
fluorescence microscopy
Protein Carbonylation
chemistry.chemical_compound
0404 agricultural biotechnology
Fluorescence microscope
Animals
protein oxidation
Sirius Red
Muscle Cells
biology
Chemistry
0402 animal and dairy science
04 agricultural and veterinary sciences
General Chemistry
carbonyl
040401 food science
040201 dairy & animal science
Primary and secondary antibodies
Fluorescence
Membrane protein
Biochemistry
Microscopy, Fluorescence
biology.protein
DNPH
Cattle
General Agricultural and Biological Sciences
Myofibril
[CHIM.OTHE]Chemical Sciences/Other
[SDV.AEN]Life Sciences [q-bio]/Food and Nutrition
Oxidation-Reduction
Intracellular
Subjects
Details
- ISSN :
- 00218561 and 15205118
- Volume :
- 55
- Issue :
- 23
- Database :
- OpenAIRE
- Journal :
- Journal of agricultural and food chemistry
- Accession number :
- edsair.doi.dedup.....e34db99ea2bac17e71a8f6318a597dc3