Phospholamban decreases the energetic efficiency of the sarcoplasmic reticulum Ca pump.
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 (45)Ca 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 (45)Ca 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 (k(leak)) was also measured at 100 nm free [Ca]. The results indicate that k(leak) 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.
Shannon
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TR
Department of Physiology, Loyola University Chicago, Maywood, Illinois 60153, USA.
Chu
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Kranias
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Bers
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DM
D000818:Animals; D002118:Calcium / Q000378:metabolism* / Q000493:pharmacokinetics; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000378:metabolism*; D007700:Kinetics; D051379:Mice; D018345:Mice, Knockout; D009206:Myocardium / Q000166:cytology* / Q000378:metabolism*; D011485:Protein Binding; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*; D019284:Thapsigargin / Q000494:pharmacology; D013816:Thermodynamics; D013997:Time Factors; D000099211:Phospholamban
D002118:Calcium; D002135:Calcium-Binding Proteins; D000252:Calcium-Transporting ATPases; D004791:Enzyme Inhibitors; D019284:Thapsigargin; D000099211:Phospholamban