Separation and identification of rat skeletal muscle proteins using two-dimensional gel electrophoresis and mass spectrometry

Skeletal muscle plays a major role in whole body protein metabolism, and ch anges in the rates of synthesis and degradation of proteins are likely to l ead to characteristic changes in the amounts of different proteins in muscl e under various physiological and pathological conditions. This paper demon strates the feasibility of a proteomic approach to analyzing the protein co mposition of skeletal muscle. We report here the initial establishment of t wo-dimensional gel electrophoresis (2-D PAGE) reference maps for mixed skel etal muscle taken from the abdominal wall of a normal adult rat We used imm obilized pH gradients of 3-10 (non-linear) and 4-7 (linear), and matrix ass isted laser desorption/ionization - time of flight mass spectrometry for pr otein identification by peptide mass fingerprinting. More than 600 protein spots were detected on each gel, of which 100 were excised and characterize d. In-gel digestion followed by peptide mass fingerprinting enabled tentati ve identification of 74 of these, which included a wide range of myofibrill ary and sarcoplasmic proteins. This database should provide the nucleus of a valuable resource for investigation of the biochemical basis of skeletal muscle pathologies in general and such specific disorders as alcoholic myop athy and injury.