ALLOSTERIC SITE STRUCTURES OF CARDIOVASCULAR CHANNELS

Summary

Principal Investigator: Arthur Karlin
Abstract: The cardiac ryanodine receptor calcium-release channel (RyR2) and the smooth-muscle large-conductance calcium-and-voltage-activated potassium (BK) channel are vital participants in normal and pathological cardiovascular physiology. Each is a tetramer of pore-forming subunits in a complex with auxiliary subunits that allosterically modulate channel behavior. FK506-binding protein (FKBP12.6) modulates RyR2, and betal modulates BK channel alpha subu'nit. In addition, these channels are allosterically modulated by ligands with therapeutic potential, such as RyR2 by JTV519. To understand the mechanisms of these allosteric interactions, the amino acid residues in the binding sites for FKBP12.6 and for JTV519 on RyR2 and for betal subunit on BK channel alpha subunit will be identified. Furthermore, changes in the contacts in different functional states of the channels will be determined. Binding-site residues in RyR2 in cardiac sarcoplasmic reticulum membrane, and in RyR2 and BK channel heterologously expressed in cultured cells, will be tagged by photoaffinity labeling, chemical crbsslinking, and foot-printing methods. The residues will be identified by protein cleavage, isolation of labeled peptides, mass spectrometry and micro-sequencing. In addition, cross-linking reactions will be directed to cysteine residues substituted by site-directed mutagenesis at specific locations on the target proteins, and cross-linking will be detected by Western blotting. Modifications of known methods are proposed to make feasible the detailed characterization of binding sites on a low abundance channel like BK and on a very large protein like the RyR2 subunit. The results will provide insights into the molecular mechanisms of allosteric interactions in these two channels and will provide molecular structural bases for cardiovascular therapeutics targeted at these channels.
Funding Period: -------------------- - --------------------
more information: NIH RePORT

Top Publications

  1. pmc Position and role of the BK channel alpha subunit S0 helix inferred from disulfide crosslinking
    Guoxia Liu
    1 Division of Cardiology, Department of Medicine, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA
    J Gen Physiol 131:537-48. 2008
  2. pmc Dysfunctional ryanodine receptors in the heart: new insights into complex cardiovascular diseases
    Steven O Marx
    Division of Cardiology, College of Physicians and Surgeons of Columbia University, New York, NY 10032, USA
    J Mol Cell Cardiol 58:225-31. 2013
  3. pmc Orientations and proximities of the extracellular ends of transmembrane helices S0 and S4 in open and closed BK potassium channels
    Xiaowei Niu
    Center for Molecular Recognition, Department of Biochemistry, College of Physicians and Surgeons, Columbia University, New York, New York, USA
    PLoS ONE 8:e58335. 2013
  4. pmc Reduced vascular smooth muscle BK channel current underlies heart failure-induced vasoconstriction in mice
    Elaine Wan
    Division of Cardiology, Columbia University College of Physicians and Surgeons, New York, NY 10032, USA
    FASEB J 27:1859-67. 2013
  5. pmc Positions of β2 and β3 subunits in the large-conductance calcium- and voltage-activated BK potassium channel
    Roland S Wu
    Division of Cardiology, Department of Medicine, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA
    J Gen Physiol 141:105-17. 2013
  6. pmc Treatment of experimental asthma using a single small molecule with anti-inflammatory and BK channel-activating properties
    Monica P Goldklang
    1S O M, Columbia University, 630 West 168th St, P and S 9 420, New York, NY 10032, USA A R M, Columbia University, Russ Berrie Medical Science Pavilion, 1150 St Nicholas Avenue, Room 520, New York, NY 10032, USA E mail
    FASEB J 27:4975-86. 2013
  7. pmc Ryanodine receptor studies using genetically engineered mice
    Alexander Kushnir
    Department of Physiology and Cellular Biophysics, Clyde and Helen Wu Center for Molecular Cardiology, New York, NY, USA
    FEBS Lett 584:1956-65. 2010
  8. pmc Locations of the beta1 transmembrane helices in the BK potassium channel
    Guoxia Liu
    Division of Cardiology, Department of Medicine, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA
    Proc Natl Acad Sci U S A 105:10727-32. 2008
  9. pmc Location of modulatory beta subunits in BK potassium channels
    Guoxia Liu
    Division of Cardiology, Department of Medicine, LaGuardia Community College, Long Island City, NY 11101, USA
    J Gen Physiol 135:449-59. 2010
  10. pmc Mice with cardiac overexpression of peroxisome proliferator-activated receptor γ have impaired repolarization and spontaneous fatal ventricular arrhythmias
    John P Morrow
    Columbia University, Division of Cardiology, PH 10 203, 622 W 168th St, New York, NY, USA
    Circulation 124:2812-21. 2011

Scientific Experts

  • Arthur Karlin
  • Roland S Wu
  • Steven O Marx
  • Guoxia Liu
  • Sergey I Zakharov
  • Takayuki Shiomi
  • Neelesh Chudasama
  • Xiaowei Niu
  • Andrew R Marks
  • Lin Yang
  • Elaine Wan
  • Monica P Goldklang
  • Jeanine D'Armiento
  • Alexander Katchman
  • Shunichi Homma
  • John P Morrow
  • Alexander Kushnir
  • David Y Chung
  • Shi Xian Deng
  • Donald W Landry
  • Jordis Trischler
  • Jared S Kushner
  • Jose F Perez-Zoghbi
  • Tina Zelonina
  • Takwi Nkyimbeng
  • Sergey Zakharov
  • JEANINE M D'ARMIENTO
  • Marc Waase
  • Xiao Wei Nui
  • Darshan Doshi
  • Shinichi Iwata
  • Christopher Kelly
  • Ira J Goldberg
  • Vaibhav Amin
  • Carolina Vasquez
  • Joshua M Lader
  • Raffay Khan
  • Gregory E Morley
  • Chad M Trent
  • Haiyan Huang
  • Ni Huiping Son
  • Xin Jin
  • Yongneng Yao
  • Howard Motoike
  • Matthew J Betzenhauser
  • Richard Weinberg
  • Robert S Kass
  • John R Bankston
  • Priscilla J Chan

Detail Information

Publications13

  1. pmc Position and role of the BK channel alpha subunit S0 helix inferred from disulfide crosslinking
    Guoxia Liu
    1 Division of Cardiology, Department of Medicine, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA
    J Gen Physiol 131:537-48. 2008
    ....
  2. pmc Dysfunctional ryanodine receptors in the heart: new insights into complex cardiovascular diseases
    Steven O Marx
    Division of Cardiology, College of Physicians and Surgeons of Columbia University, New York, NY 10032, USA
    J Mol Cell Cardiol 58:225-31. 2013
    ..Correcting these defects using either genetic manipulation (knock-in) in mice, or specific and novel small molecules ameliorates the RyR2 dysfunction, reducing the progression to heart failure and the incidence of arrhythmias...
  3. pmc Orientations and proximities of the extracellular ends of transmembrane helices S0 and S4 in open and closed BK potassium channels
    Xiaowei Niu
    Center for Molecular Recognition, Department of Biochemistry, College of Physicians and Surgeons, Columbia University, New York, New York, USA
    PLoS ONE 8:e58335. 2013
    ....
  4. pmc Reduced vascular smooth muscle BK channel current underlies heart failure-induced vasoconstriction in mice
    Elaine Wan
    Division of Cardiology, Columbia University College of Physicians and Surgeons, New York, NY 10032, USA
    FASEB J 27:1859-67. 2013
    ..These findings identify electrical dysregulation within VSM, specifically the reduction of BK currents, as a key molecular mechanism sensitizing resistance vessels to pressure-induced vasoconstriction in systolic HF...
  5. pmc Positions of β2 and β3 subunits in the large-conductance calcium- and voltage-activated BK potassium channel
    Roland S Wu
    Division of Cardiology, Department of Medicine, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA
    J Gen Physiol 141:105-17. 2013
    ..In no case was N-type inactivation by the N-terminal tail of mβ3 perturbed. Although the mβ3 loop can move, its position with Cys152 near αK296, in which it blocks IbTX binding, is likely favored...
  6. pmc Treatment of experimental asthma using a single small molecule with anti-inflammatory and BK channel-activating properties
    Monica P Goldklang
    1S O M, Columbia University, 630 West 168th St, P and S 9 420, New York, NY 10032, USA A R M, Columbia University, Russ Berrie Medical Science Pavilion, 1150 St Nicholas Avenue, Room 520, New York, NY 10032, USA E mail
    FASEB J 27:4975-86. 2013
    ..These findings suggest that rottlerin, with both anti-inflammatory and ASM relaxation properties, may have benefit in treating asthma...
  7. pmc Ryanodine receptor studies using genetically engineered mice
    Alexander Kushnir
    Department of Physiology and Cellular Biophysics, Clyde and Helen Wu Center for Molecular Cardiology, New York, NY, USA
    FEBS Lett 584:1956-65. 2010
    ..In this review we summarize the RyR mouse models and how they have enhanced our understanding of the RyR channels and their roles in cellular physiology and disease...
  8. pmc Locations of the beta1 transmembrane helices in the BK potassium channel
    Guoxia Liu
    Division of Cardiology, Department of Medicine, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA
    Proc Natl Acad Sci U S A 105:10727-32. 2008
    ..TM1 and TM2 are in position to contribute, in concert with the extracellular loop and the intracellular N- and C-terminal tails of beta1, to the modulation of BK channel function...
  9. pmc Location of modulatory beta subunits in BK potassium channels
    Guoxia Liu
    Division of Cardiology, Department of Medicine, LaGuardia Community College, Long Island City, NY 11101, USA
    J Gen Physiol 135:449-59. 2010
    ..Thus, each beta lies between and can interact with the voltage-sensing domains of two adjacent alpha subunits...
  10. pmc Mice with cardiac overexpression of peroxisome proliferator-activated receptor γ have impaired repolarization and spontaneous fatal ventricular arrhythmias
    John P Morrow
    Columbia University, Division of Cardiology, PH 10 203, 622 W 168th St, New York, NY, USA
    Circulation 124:2812-21. 2011
    ..It is difficult to distinguish the contribution of cardiomyocyte lipid accumulation from the contribution of global metabolic defects to the increased incidence of sudden death and electric abnormalities...
  11. pmc Location of the beta 4 transmembrane helices in the BK potassium channel
    Roland S Wu
    Division of Cardiology, Department of Medicine, College of Physicians and Surgeons, Columbia University, New York, New York 10032, USA
    J Neurosci 29:8321-8. 2009
    ..That disulfide crosslinking caused only small functional perturbations is consistent with the proximity of the extracellular ends of TM2 to S0 and of TM1 to S1 and to S2, in both the open and closed states...
  12. pmc Location of KCNE1 relative to KCNQ1 in the I(KS) potassium channel by disulfide cross-linking of substituted cysteines
    David Y Chung
    Department of Pharmacology, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA
    Proc Natl Acad Sci U S A 106:743-8. 2009
    ..Thus, the first E1 orientation strongly favors the open state, while the approximately opposite orientation favors the closed state...
  13. ncbi The BK potassium channel in the vascular smooth muscle and kidney: α- and β-subunits
    Roland S Wu
    Division of Cardiology, Department of Medicine, College of Physicians and Surgeons, Columbia University, New York, New York, USA
    Kidney Int 78:963-74. 2010
    ..In this review, we will focus on function, regulation, and clinical significance of the BK channel. We will also specifically concentrate on one of its β-subunits, β1, which is important in vascular and renal physiology...