Activity of creatine kinase in a contracting mammalian muscle of uniform fiber type.
We investigated whether the creatine kinase-catalyzed phosphate exchange between PCr and gamma ATP in vivo equilibrated with cellular substrates and products as predicted by in vitro kinetic properties of the enzyme, or was a function of ATPase activity as predicted by obligatory "creatine phosphate shuttle" concepts. A transient NMR spin-transfer method was developed, tested, and applied to resting and stimulated ex vivo muscle, the soleus, which is a cellularly homogeneous slow-twitch mammalian muscle, to measure creatine kinase kinetics. The forward and reverse unidirectional CK fluxes were equal, being 1.6 mM.s-1 in unstimulated muscle at 22 degrees C, and 2.7 mM.s-1 at 30 degrees C. The CK fluxes did not differ during steady-state stimulation conditions giving a 10-fold range of ATPase rates in which the ATP/PCr ratio increased from approximately 0.3 to 1.6. The observed kinetic behavior of CK activity in the muscle was that expected from the enzyme in vitro in a homogeneous solution only if account was taken of inhibition by an anion-stabilized quaternary dead-end enzyme complex: E.Cr.MgADP.anion. The CK fluxes in soleus were not a function of ATPase activity as predicted by obligatory phosphocreatine shuttle models for cellular energetics.
Department of Chemical Engineering, University of California at Santa Barbara 93106.
E W
EW
McFarland
M J
MJ
Kushmerick
T S
TS
Moerland
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D055592:Biophysical Phenomena; D001703:Biophysics; D002415:Cats; D003402:Creatine Kinase / Q000378:metabolism*; D066298:In Vitro Techniques; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008954:Models, Biological; D009119:Muscle Contraction / Q000502:physiology; D018482:Muscle, Skeletal / Q000201:enzymology* / Q000502:physiology; D010477:Perfusion; D010725:Phosphocreatine / Q000378:metabolism
D010725:Phosphocreatine; D000255:Adenosine Triphosphate; D003402:Creatine Kinase; D000251:Adenosine Triphosphatases