JBC

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Originally published In Press as doi:10.1074/jbc.M208124200 on September 24, 2002

J. Biol. Chem., Vol. 277, Issue 48, 46712-46719, November 29, 2002
This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
277/48/46712    most recent
M208124200v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Liu, Z.
Right arrow Articles by Wagenknecht, T.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Liu, Z.
Right arrow Articles by Wagenknecht, T.
Social Bookmarking
Add to CiteULike   Add to Complore   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati  
What's this?

Three-dimensional Reconstruction of the Recombinant Type 2 Ryanodine Receptor and Localization of Its Divergent Region 1*

Zheng LiuDagger §, Jing Zhang, Pin Li, S. R. Wayne Chen, and Terence WagenknechtDagger ||

From the Dagger  Wadsworth Center, New York State Department of Health, Albany, New York 12201, the  Department of Physiology and Biophysics, Biochemistry, and Molecular Biology, University of Calgary, Calgary, Alberta T2N 4N1, Canada, and the || Department of Biomedical Sciences, School of Public Health, State University of New York at Albany, Albany, New York 12201

Isoform 2 of the ryanodine receptor (RyR2) is the major calcium release channel in cardiac muscle. In the present study, two kinds of RyR2 cDNA were constructed, one encoding the wild type mouse RyR2 (RyR2wt) and the other encoding modified RyR2, into which was inserted a cDNA encoding green fluorescent protein (GFP). GFP was inserted into the divergent region 1 (DR1) of RyR2, after the Asp-4365 (RyR2D4365-GFP). HEK293 cells expressing both RyR2wt and RyR2D4365-GFP cDNAs showed caffeine- and ryanodine-sensitive calcium release, demonstrating that both wild type and modified RyR2s form functional calcium release channels. Cells expressing the fusion protein, RyR2D4365-GFP, were readily identified by their fluorescence due to the presence of GFP, indicating that the inserted GFP folded properly. Both expressed RyR2s were purified from cell lysates in a single step by affinity chromatography using a GST-FKBP12.6 as the affinity ligand. Cryoelectron microscopy of purified RyR2s showed structurally intact receptors, and three-dimensional reconstructions were obtained by single particle image processing. The three-dimensional reconstruction of RyR2wt appeared very similar to that of the native RyR2 purified from dog heart. The location of the inserted GFP, and consequently of DR1, was mapped on the three-dimensional structure of RyR2 to one of the subunit's characteristic domains, domain 3, also known as the "handle" domain. This study describes the first internal fusion of a protein into a ryanodine receptor, and it demonstrates the potential of this technology for localizing functional and structural domains on the three-dimensional structure of RyR.

* This work was supported by the Muscular Dystrophy Association and National Institutes of Health Grant AR40615 (to T. W.) and by research grants from the Canadian Institutes of Health Research and the Heart and Stroke Foundation of Alberta, Northwest Territory, and Nunavut (to S. R. W. C.).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.

§ To whom correspondence and reprint requests should be addressed: Wadsworth Center, New York State Department of Health, Albany, NY 12201-0509. Tel.: 518-474-7895; Fax: 518-474-7992; E-mail: liuz@wadsworth.org.

Copyright © 2002 by The American Society for Biochemistry and Molecular Biology, Inc.
Add to CiteULike CiteULike   Add to Complore Complore   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Reddit Reddit   Add to Technorati Technorati    What's this?


This article has been cited by other articles:


Home page
 Cardiovasc ResHome page
C. H. George
Sarcoplasmic reticulum Ca2+ leak in heart failure: mere observation or functional relevance?
Cardiovasc Res, January 15, 2008; 77(2): 302 - 314.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
X. Meng, B. Xiao, S. Cai, X. Huang, F. Li, J. Bolstad, R. Trujillo, J. Airey, S. R. W. Chen, T. Wagenknecht, et al.
Three-Dimensional Localization of Serine 2808, a Phosphorylation Site in Cardiac Ryanodine Receptor
J. Biol. Chem., August 31, 2007; 282(35): 25929 - 25939.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
R. Wang, W. Chen, S. Cai, J. Zhang, J. Bolstad, T. Wagenknecht, Z. Liu, and S. R. W. Chen
Localization of an NH2-terminal Disease-causing Mutation Hot Spot to the "Clamp" Region in the Three-dimensional Structure of the Cardiac Ryanodine Receptor
J. Biol. Chem., June 15, 2007; 282(24): 17785 - 17793.
[Abstract] [Full Text] [PDF]

Home page
 Biophys. JHome page
A. J. Tanskanen, J. L. Greenstein, A. Chen, S. X. Sun, and R. L. Winslow
Protein Geometry and Placement in the Cardiac Dyad Influence Macroscopic Properties of Calcium-Induced Calcium Release
Biophys. J., May 15, 2007; 92(10): 3379 - 3396.
[Abstract] [Full Text] [PDF]

Home page
 Physiol. Rev.Home page
H. E. D. J. ter Keurs and P. A. Boyden
Calcium and Arrhythmogenesis
Physiol Rev, April 1, 2007; 87(2): 457 - 506.
[Abstract] [Full Text] [PDF]

Home page
 Proc. Natl. Acad. Sci. USAHome page
D. C. Sheridan, H. Takekura, C. Franzini-Armstrong, K. G. Beam, P. D. Allen, and C. F. Perez
Bidirectional signaling between calcium channels of skeletal muscle requires multiple direct and indirect interactions
PNAS, December 26, 2006; 103(52): 19760 - 19765.
[Abstract] [Full Text] [PDF]

Home page
 Ann. N. Y. Acad. Sci.Home page
R. L WINSLOW, A. TANSKANEN, M. CHEN, and J. L GREENSTEIN
Multiscale Modeling of Calcium Signaling in the Cardiac Dyad
Ann. N.Y. Acad. Sci., October 1, 2006; 1080(1): 362 - 375.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
Z. Liu, R. Wang, J. Zhang, S. R. W. Chen, and T. Wagenknecht
Localization of a Disease-associated Mutation Site in the Three-dimensional Structure of the Cardiac Muscle Ryanodine Receptor
J. Biol. Chem., November 11, 2005; 280(45): 37941 - 37947.
[Abstract] [Full Text] [PDF]

Home page
 Biophys. JHome page
L. Carbonneau, D. Bhattacharya, D. C. Sheridan, and R. Coronado
Multiple Loops of the Dihydropyridine Receptor Pore Subunit Are Required for Full-Scale Excitation-Contraction Coupling in Skeletal Muscle
Biophys. J., July 1, 2005; 89(1): 243 - 255.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
S. Papadopoulos, V. Leuranguer, R. A. Bannister, and K. G. Beam
Mapping Sites of Potential Proximity between the Dihydropyridine Receptor and RyR1 in Muscle Using a Cyan Fluorescent Protein-Yellow Fluorescent Protein Tandem as a Fluorescence Resonance Energy Transfer Probe
J. Biol. Chem., October 15, 2004; 279(42): 44046 - 44056.
[Abstract] [Full Text] [PDF]

Home page
 Biophys. JHome page
S. Peng, N. G. Publicover, G. J. Kargacin, D. Duan, J. A. Airey, and J. L. Sutko
Imaging Single Cardiac Ryanodine Receptor Ca2+ Fluxes in Lipid Bilayers
Biophys. J., January 1, 2004; 86(1): 134 - 144.
[Abstract] [Full Text] [PDF]

Home page
 Biophys. JHome page
S. Peng, N. G. Publicover, J. A. Airey, J. E. Hall, H. T. Haigler, D. Jiang, S. R. W. Chen, and J. L. Sutko
Diffusion of Single Cardiac Ryanodine Receptors in Lipid Bilayers Is Decreased by Annexin 12
Biophys. J., January 1, 2004; 86(1): 145 - 151.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
J. Zhang, Z. Liu, H. Masumiya, R. Wang, D. Jiang, F. Li, T. Wagenknecht, and S. R. W. Chen
Three-dimensional Localization of Divergent Region 3 of the Ryanodine Receptor to the Clamp-shaped Structures Adjacent to the FKBP Binding Sites
J. Biol. Chem., April 11, 2003; 278(16): 14211 - 14218.
[Abstract] [Full Text] [PDF]

Home page
 Sci SignalHome page
E. M. Adler, N. R. Gough, and L. B. Ray
2002: Signaling Breakthroughs of the Year
Sci. Signal., January 7, 2003; 2003(164): eg1 - eg1.
[Full Text] [PDF]


HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
 All ASBMB Journals   Molecular and Cellular Proteomics 
 Journal of Lipid Research   ASBMB Today 
Copyright © 2002 by the American Society for Biochemistry and Molecular Biology.