CHANNELS, RECEPTORS, AND ELECTRICAL SIGNALING |
THREE-DIMENSIONAL VISUALIZATION OF FKBP12.6 BINDING TO
AN OPEN CONFORMATION OF CARDIAC RYANODINE RECEPTOR
Manjuli Rani Sharma 1*, Loice H Jeyakumar 2, Sidney Fleischer 3 and Terence Wagenknecht 1
1 Wadsworth Center, New York State Department of Health
2 Dept. of Medicine, Vanderbilt University of Medical Center
3 Department of Biological Sciences, Vanderbilt University
* To whom correspondence should be addressed. E-mail: manjuli{at}wadsworth.org.
Submitted on March 25, 2005
Revised on April 18, 2005
Accepted on 20 September 2005
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Abstract |
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The cardiac isoform of the ryanodine receptor (RyR2) from dog binds predominantly a 12.6-kDa isoform of the FK506-binding protein (FKBP12.6), whereas RyR2 from other species binds both FKBP12.6 and the closely related isoform FKBP12. The role played by FKBP12.6 in modulating calcium release by RyR2 is unclear at present. We have used cryo-electron microscopy and 3D reconstruction techniques to determine the binding position of FKBP12.6 on the surface of canine RyR2. Buffer conditions that should favor the open state of RyR2 were used. Quantitative comparison of 3D reconstructions of RyR2 in the presence and absence of FKBP12.6 reveals that FKBP12.6 binds along the sides of the square-shaped cytoplasmic region of the receptor, adjacent to domain 9, which forms part of the four the clamp (corner-forming) structures. The location of the FKBP12.6 binding site on "open" RyR2 appears similar, but slightly displaced (by 1-2 nm) from that found previously for FKBP12 binding to the skeletal muscle ryanodine receptor that was in buffer that favors the "closed" state. The conformation of RyR2 containing bound FKBP12.6 differs considerably from that in the absence of FKBP12.6, particularly in the transmembrane region and in the clamp structures. The X-ray structure of FKBP12.6 was docked into the region of the 3D reconstruction that is attributable to bound FKBP12.6, to show the relative orientations of amino-acid residues (Gln-31, Asn-32, Phe-59) that have been implicated as being critical in interactions with RyR2. A thorough understanding of the structural basis of RyR2-FKBP12.6 interaction should aid in understanding the roles that have been proposed for FKBP12.6 in heart failure and in certain forms of sudden cardiac death.
Key Words:
Cryo-EM: cryo-electron microscopy, FKBP12.6 : 12.6 isoform of FK506-binding protein, RyR1: skeletal muscle ryanodine receptor, RyR2: cardiac muscle ryanodine receptor, RyR: ryanodine receptor, SR: Sarcoplasmic Reticulum
