Functional coupling between sarcoplasmic-reticulum-bound creatine kinase and Ca2+-ATPase

Authors


Correspondence to P. Korge, Dept VCAPP, Washington State University, Pullman WA 99164, USA
Fax:+1 509 335 4650.

Abstract

We investigated the role of creatine kinase bound to sarcoplasmic reticulum membranes of fast skeletal muscle in the local regeneration of ATP and the possible physiological significance of this regeneration for calcium pump function. Our results indicate that ADP produced by sarcoplasmic reticulum Ca2+-ATPase is effectively phosphorylated by creatine kinase in the presence of creatine phosphate. This phosphorylation is an important function of the membrane-bound creatine kinase because accumulation of ADP has a depressive effect on Ca2+-uptake by sarcoplasmic reticulum vesicles. The concentration-dependent depression of Ca2+-uptake by ADP was especially pronounced when there was strong back inhibition by high intravesicular [Ca2+]. ATP regenerated by endogenous creatine kinase was not in free equilibrium with the ATP in the surrounding medium, but was used preferentially by Ca2+-ATPase for Ca2+-uptake. Efficient translocation of ATP from creatine kinase to Ca2+-ATPase, despite the presence of an ATP trap in the surrounding medium, can be explained by close localization of creatine kinase and Ca2+-ATPase on the sarcoplasmic reticulum membranes. These results suggest the existence of functional coupling between creatine kinase and Ca2+-ATPase on skeletal muscle sarcoplasmic reticulum membranes. Several factors (amount of membrane-bound creatine kinase, oxidation of SH groups of creatine kinase, decrease in [phosphocreatine]) can influence the ability of creatine kinase/phosphocreatine system to support a low ADP/ATP ratio and fuel the Ca2+-pump with ATP. These factors may become operative in the living cells, influencing functional coupling between creatine kinase and Ca2+-ATPase and may have an indirect effect on Ca2+-pump function before Ca2+-ATPase itself is affected.

Abbreviation
SR

sarcoplasmic reticulum

P-creatine

phosphocreatine

P-pyruvate

phospho(enol)pyruvate

Ap5A

adenosine(5′)pentaphospho(5′)adenosine

Dnp-F

1-fluoro-2,4-dinitrobenzene

MalNEt

N-ethylmaleimide

Enzyme
 

Ca2+-ATPase (EC 3.6.1.38)

 

creatine kinase (EC 2.7.3.2)

 

pyruvate kinase (EC 2.7.1.40)

 

hexokinase (EC 2.7.1.1)

 

adenylate kinase (EC 2.7.4.3)

 

lactic dehydrogenase (EC 1.1.1.27)

 

glucose-6-phosphate dehydrogenase (EC 1.1.1.49)

 

apyrase (EC 3.6.1.5)

 

glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.12)

Ancillary