SARCOLEMMAL CARBONIC-ANHYDRASE IN RED AND WHITE-RABBIT SKELETAL-MUSCLE

Sarcolemmal vesicles of white and red skeletal muscles of the rabbit were prepared by consecutive density gradient centrifugations in sucrose and dextran according to Seiler and Fleischer (1982, J. Biol. Chem. 257, 13,862-13,871). White and red muscle membrane fractions enriched in sarcolemma were characterized by high ouabain-sensitive Na+, K+-ATPase, by high Mg2+-ATPase activity, and by a high cholesterol content. Ca2+-ATPase activity, a marker enzyme for sarcoplasmic reticulum, was not detectable in the highly purified white and red muscle sarcolemmal fractions. White and red muscle sarcolemmal fractions exhibited no significant differences with regard to Na+, K+-ATPase, Mg2+-ATPase, and cholesterol. Specific activity of carbonic anhydrase in white muscle sarcolemmal fractions was 38 U · ml/mg and was 17.6 U · ml/mg in red muscle sarcolemma. Inhibition properties of sarcolemmal carbonic anhydrase were analyzed for acetazolamide, chlorzolamide, and cyanate. White muscle sarcolemmal carbonic anhydrase is characterized by inhibition constants, KI, toward acetazolamide of 4.6 · 10-8 m, toward chlorzolamide of 0.75 · 10-8 m, and toward cyanate of 1.3 · 10-4 m. Red muscle sarcolemmal carbonic anhydrase is characterized by KI values toward acetazolamide of 8.1 · 10-8 m, toward chlorzolamide of 6.3 · 10-8 m, and toward cyanate of 0.81 · 10-4 m. In contrast to the high specific carbonic anhydrase activities in sarcolemma, carbonic anhydrase activity in sarcoplasmic reticulum from white muscle varied between values of only 0.7 and 3.3 U · ml/mg. Carbonic anhydrase of red muscle sarcoplasmic reticulum ranged from 2.4 to 3.7 U · ml/mg. © 1990.