Separation of cardiac myosin heavy chains by gradient SDS-PAGE.
Separation of alpha- and beta-myosin heavy chains (MHCs) in cardiac ventricles of rats by gradient sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was accomplished and compared with the separation of myosin isozymes obtained with pyrophosphate gels. Whole muscle homogenates were electrophoresed on a 4-9% linear gradient SDS polyacrylamide gel for 3-4 h. MHC bands were identified by the migration distance relative to a MHC standard and immunoblot results with a monoclonal antibody to MHC. The MHC bands were further identified as alpha and beta based on the electrophoretic mobility of ventricular homogenates from hypothyroid and hyperthyroid rats and ventricular and slow soleus skeletal muscle homogenates from control rats. The beta-MHC migrated faster than alpha-MHC, and laser densitometry revealed separate peaks when both MHCs were present. With homogenates containing MHC ranging from 0 to 100% alpha, the separation of MHCs with gradient SDS-PAGE correlated highly (r = 0.97) with separation of myosin isozymes by pyrophosphate gel electrophoresis. The SDS-PAGE technique reported herein is a quick, valid, and direct method for the identification and quantification of ventricular MHCs.
Department of Kinesiology, University of Michigan, Ann Arbor 48109-2214.
K A
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Esser
M O
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Boluyt
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White
D000818:Animals; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D006352:Heart Ventricles / Q000378:metabolism; D006980:Hyperthyroidism / Q000378:metabolism; D007037:Hypothyroidism / Q000378:metabolism; D008297:Male; D008970:Molecular Weight; D009206:Myocardium / Q000378:metabolism*; D015879:Myosin Subfragments; D009218:Myosins / Q000302:isolation & purification*; D010446:Peptide Fragments / Q000302:isolation & purification*; D051381:Rats; D011919:Rats, Inbred Strains; D012016:Reference Values
D015879:Myosin Subfragments; D010446:Peptide Fragments; D009218:Myosins