title: Protein composition and function of red and white skeletal muscle mitochondria.
abstract: Red and white muscles are faced with very different energetic demands. However, it is unclear whether relative mitochondrial protein expression is different between muscle types. Mitochondria from red and white porcine skeletal muscle were isolated with a Percoll gradient. Differences in protein composition were determined using blue native (BN)-PAGE, two-dimensional differential in gel electrophoresis (2D DIGE), optical spectroscopy, and isobaric tag for relative and absolute quantitation (iTRAQ). Complex IV and V activities were compared using BN-PAGE in-gel activity assays, and maximal mitochondrial respiration rates were assessed using pyruvate (P) + malate (M), glutamate (G) + M, and palmitoyl-carnitine (PC) + M. Without the Percoll step, major cytosolic protein contamination was noted for white mitochondria. Upon removal of contamination, very few protein differences were observed between red and white mitochondria. BN-PAGE showed no differences in the subunit composition of Complexes I-V or the activities of Complexes IV and V. iTRAQ analysis detected 358 mitochondrial proteins, 69 statistically different. Physiological significance may be lower: at a 25% difference, 48 proteins were detected; at 50%, 14 proteins were detected; and 3 proteins were detected at a 100%. Thus any changes could be argued to be physiologically modest. One area of difference was fat metabolism where four β-oxidation enzymes were ∼25% higher in red mitochondria. This was correlated with a 40% higher rate of PC+M oxidation in red mitochondria compared with white mitochondria with no differences in P+M and G+M oxidation. These data suggest that metabolic demand differences between red and white muscle fibers are primarily matched by the number of mitochondria and not by significant alterations in the mitochondria themselves.
authors.affiliation: National Heart, Lung, and Blood Institute/NIH, 10 Center Drive, Bethesda, MD 20892, USA. glancybp@nhlbi.nih.gov
authors.forename: Brian
authors.initials: B
authors.lastname: Glancy
authors.forename: Robert S
authors.initials: RS
authors.lastname: Balaban
mesh_terms: D000818:Animals; D006801:Humans; D008931:Mitochondria, Muscle / Q000737:chemistry* / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000737:chemistry / Q000378:metabolism*; D018656:Muscle Fibers, Fast-Twitch / Q000737:chemistry / Q000166:cytology*; D018657:Muscle Fibers, Slow-Twitch / Q000737:chemistry / Q000166:cytology*; D018482:Muscle, Skeletal / Q000737:chemistry* / Q000166:cytology* / Q000378:metabolism; D010084:Oxidation-Reduction; D010085:Oxidative Phosphorylation
chemical_list: D024101:Mitochondrial Proteins