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J. Biol. Chem., Vol. 276, Issue 10, 7195-7201, March 9, 2001
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From the We tested the hypothesis that increased
Sarcoplasmic reticulum (SR) Ca content
([Ca]SRT) in phospholamban knockout mice (PLB-KO) is because of increased SR Ca pump efficiency defined by the steady-state SR [Ca] gradient. The time course of
thapsigargin-sensitive ATP-dependent
45Ca influx into and efflux out of cardiac SR vesicles from
PLB-KO and wild-type (WT) mice was measured at 100 nM free
[Ca]. We found that PLB decreased the initial SR Ca uptake rate (0.13 versus 0.31 nmol/mg/s) and decreased steady-state
45Ca content (0.9 versus 4.1 nmol/mg protein).
Furthermore, at similar total SR [Ca], the pump-mediated Ca efflux
rate was higher in WT (0.065 versus 0.037 nmol/mg/s). The
pump-independent leak rate constant
(kleak) was also measured at 100 nM free [Ca]. The results indicate that
kleak was <1% of pump-mediated
backflux and was not different among nonpentameric mutant PLB
(PLB-C41F), WT pentameric PLB (same expression level), and PLB-KO.
Therefore differences in passive SR Ca leak cannot be the cause of the
higher thapsigargin-sensitive Ca efflux from the WT
membranes. We conclude that the decreased total SR [Ca] in WT mice is
caused by decreased SR Ca influx rate, an increased Ca-pump backflux,
and unaltered leak. Based upon both thermodynamic and kinetic analysis,
we conclude that PLB decreases the energetic efficiency of the SR Ca pump.
Phospholamban Decreases the Energetic Efficiency of the
Sarcoplasmic Reticulum Ca Pump*
,¶
Department of Physiology, Loyola
University Chicago, Maywood, Illinois 60153 and the
§ Department of Pharmacology and Cell Biophysics, University
of Cincinnati, Cincinnati, Ohio 45267
*
This work was supported by National Institutes of Health
Grants HL30077 and HL09412 and by American Heart Association Grant 9804712X.The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
Copyright © 2001 by The American Society for Biochemistry and Molecular Biology, Inc.
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