Activation of sarcoplasmic reticulum Ca(2+)-ATPase by Mn2+: a Mn2+ binding study.
We used Mn2+ as an analogue for Mg2+ to examine the minimum requirement of divalent cations for the rapid turnover of the sarcoplasmic reticulum ATPase. We measured the binding of Ca2+ and Mn2+ to the purified Ca(2+)-ATPase during steady-state hydrolysis of MnATP at 2 degrees C and pH 7. In the presence of 20 microM Ca2+, Mn2+ was as effective as Mg2+ in stimulating ATPase activity and the maximal activation of ATP hydrolysis was observed at 0.1 mM MnCl2. Under these conditions, 2 mol of Ca2+ were bound per mol of the ADP-sensitive phosphoenzyme, whereas no Ca2+ was bound to the ADP-insensitive phosphoenzyme. On the other hand, the stoichiometry for ATP-dependent binding of Mn2+ to these intermediates was about 1. We found that Mn2+ remained bound to the ADP-insensitive phosphoenzyme even in the presence of added chelator. In the absence of ATP, we detected a low level of Mn2+ binding, which reached 0.4 mol per mol of the phosphorylation site at 0.1 mM free Mn2+. We present evidence that this extra Mn2+ binding did not affect the rate of decomposition of the ADP-sensitive phosphoenzyme, which was the rate-limiting step for ATP hydrolysis under the conditions used.(ABSTRACT TRUNCATED AT 250 WORDS)
Ogurusu
T
T
Department of Molecular Physiology, National Cardiovascular Center Research Institute, Osaka.
Wakabayashi
S
S
Shigekawa
M
M
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D001692:Biological Transport; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D002712:Chlorides; D004789:Enzyme Activation; D006868:Hydrolysis; D015636:Magnesium Chloride / Q000378:metabolism; D008345:Manganese / Q000378:metabolism*; D017895:Manganese Compounds; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D013379:Substrate Specificity
D002712:Chlorides; D017895:Manganese Compounds; D015636:Magnesium Chloride; D008345:Manganese; D000255:Adenosine Triphosphate; D000252:Calcium-Transporting ATPases; C025340:manganese chloride