Reversible Covalent Binding to Cardiac Troponin C by the Ca2+-Sensitizer Levosimendan.
The binding of Ca2+to cardiac troponin C (cTnC) triggers contraction in heart muscle. In the diseased heart, the myocardium is often desensitized to Ca2+, which leads to impaired contractility. Therefore, compounds that sensitize cardiac muscle to Ca2+(Ca2+-sensitizers) have therapeutic promise. The only Ca2+-sensitizer used regularly in clinical settings is levosimendan. While the primary target of levosimendan is thought to be cTnC, the molecular details of this interaction are not well understood. In this study, we used mass spectrometry, computational chemistry, and nuclear magnetic resonance spectroscopy to demonstrate that levosimendan reacts specifically with cysteine 84 of cTnC to form a reversible thioimidate bond. We also showed that levosimendan only reacts with the active, Ca2+-bound conformation of cTnC. Finally, we propose a structural model of levosimendan bound to cTnC, which suggests that the Ca2+-sensitizing function of levosimendan is due to stabilization of the Ca2+-bound conformation of cTnC.
Randall Division of Cell and Molecular Biophysics and British Heart Foundation Centre of Research Excellence, King's College London , New Hunt's House, Guy's Campus, London, SE1 1UL, U.K.|Department of Biochemistry, Faculty of Medicine and Dentistry, University of Alberta , Edmonton, Alberta T6G 2H7, Canada.
Ian M
IM
Robertson
Department of Biochemistry, Faculty of Medicine and Dentistry, University of Alberta , Edmonton, Alberta T6G 2H7, Canada.
Sandra E
SE
Pineda-Sanabria
Randall Division of Cell and Molecular Biophysics and British Heart Foundation Centre of Research Excellence, King's College London , New Hunt's House, Guy's Campus, London, SE1 1UL, U.K.
Ziqian
Z
Yan
Randall Division of Cell and Molecular Biophysics and British Heart Foundation Centre of Research Excellence, King's College London , New Hunt's House, Guy's Campus, London, SE1 1UL, U.K.
Thomas
T
Kampourakis
Randall Division of Cell and Molecular Biophysics and British Heart Foundation Centre of Research Excellence, King's College London , New Hunt's House, Guy's Campus, London, SE1 1UL, U.K.
Yin-Biao
YB
Sun
Department of Biochemistry, Faculty of Medicine and Dentistry, University of Alberta , Edmonton, Alberta T6G 2H7, Canada.
Brian D
BD
Sykes
Randall Division of Cell and Molecular Biophysics and British Heart Foundation Centre of Research Excellence, King's College London , New Hunt's House, Guy's Campus, London, SE1 1UL, U.K.
Malcolm
M
Irving
D002118:Calcium / Q000378:metabolism*; D066241:Carbon-13 Magnetic Resonance Spectroscopy; D002316:Cardiotonic Agents / Q000378:metabolism*; D006835:Hydrazones / Q000378:metabolism*; D009206:Myocardium / Q000378:metabolism*; D011485:Protein Binding; D011724:Pyridazines / Q000378:metabolism*; D000077464:Simendan; D019209:Troponin C / Q000378:metabolism*
D002316:Cardiotonic Agents; D006835:Hydrazones; D011724:Pyridazines; D019209:Troponin C; D000077464:Simendan; D002118:Calcium