Note to users. If you're seeing this message, it means that your browser cannot find this page's style/presentation instructions -- or possibly that you are using a browser that does not support current Web standards. Find out more about why this message is appearing, and what you can do to make your experience of our site the best it can be.
Subscribe

Logo for

Science 323 (5914): 633-637

Copyright © 2009 by the American Association for the Advancement of Science

Ligand-Dependent Equilibrium Fluctuations of Single Calmodulin Molecules

Jan Philipp Junker,1 Fabian Ziegler,1 Matthias Rief1,2*

Abstract: Single-molecule force spectroscopy allows superb mechanical control of protein conformation. We used a custom-built low-drift atomic force microscope to observe mechanically induced conformational equilibrium fluctuations of single molecules of the eukaryotic calcium-dependent signal transducer calmodulin (CaM). From this data, the ligand dependence of the full energy landscape can be reconstructed. We find that calcium ions affect the folding kinetics of the individual CaM domains, whereas target peptides stabilize the already folded structure. Single-molecule data of full length CaM reveal that a wasp venom peptide binds noncooperatively to CaM with 2:1 stoichiometry, whereas a target enzyme peptide binds cooperatively with 1:1 stoichiometry. If mechanical load is applied directly to the target peptide, real-time binding/unbinding transitions can be observed.

1 Physik Department E22, Technische Universität München, James-Franck-Strasse, 85748 München, Germany.
2 Munich Center for Integrated Protein Science, 81377 München, Germany.

* To whom correspondence should be addressed. E-mail: mrief{at}ph.tum.de

THIS ARTICLE HAS BEEN CITED BY OTHER ARTICLES:
Protein Folding Under Mechanical Forces: A Physiological View.
Y. Javadi, J. M. Fernandez, and R. Perez-Jimenez (2013)
Physiology 28, 9-17
   Abstract »    Full Text »    PDF »
Single-molecule dissection of the high-affinity cohesin-dockerin complex.
S. W. Stahl, M. A. Nash, D. B. Fried, M. Slutzki, Y. Barak, E. A. Bayer, and H. E. Gaub (2012)
PNAS 109, 20431-20436
   Abstract »    Full Text »    PDF »
Calcium-dependent folding of single calmodulin molecules.
J. Stigler and M. Rief (2012)
PNAS 109, 17814-17819
   Abstract »    Full Text »    PDF »
Counteracting chemical chaperone effects on the single-molecule {alpha}-synuclein structural landscape.
A. C. M. Ferreon, M. M. Moosa, Y. Gambin, and A. A. Deniz (2012)
PNAS 109, 17826-17831
   Abstract »    Full Text »    PDF »
Single Reconstituted Neuronal SNARE Complexes Zipper in Three Distinct Stages.
Y. Gao, S. Zorman, G. Gundersen, Z. Xi, L. Ma, G. Sirinakis, J. E. Rothman, and Y. Zhang (2012)
Science 337, 1340-1343
   Abstract »    Full Text »    PDF »
Energy landscape analysis of native folding of the prion protein yields the diffusion constant, transition path time, and rates.
H. Yu, A. N. Gupta, X. Liu, K. Neupane, A. M. Brigley, I. Sosova, and M. T. Woodside (2012)
PNAS 109, 14452-14457
   Abstract »    Full Text »    PDF »
Alternative Pathways for Association and Dissociation of the Calmodulin-binding Domain of Plasma Membrane Ca2+-ATPase Isoform 4b (PMCA4b).
J. T. Penniston, A. J. Caride, and E. E. Strehler (2012)
J. Biol. Chem. 287, 29664-29671
   Abstract »    Full Text »    PDF »
Mutation of Conserved Histidines Alters Tertiary Structure and Nanomechanics of Consensus Ankyrin Repeats.
W. Lee, J. Strumpfer, V. Bennett, K. Schulten, and P. E. Marszalek (2012)
J. Biol. Chem. 287, 19115-19121
   Abstract »    Full Text »    PDF »
The Complex Folding Network of Single Calmodulin Molecules.
J. Stigler, F. Ziegler, A. Gieseke, J. C. M. Gebhardt, and M. Rief (2011)
Science 334, 512-516
   Abstract »    Full Text »    PDF »
Fast-folding {alpha}-helices as reversible strain absorbers in the muscle protein myomesin.
F. Berkemeier, M. Bertz, S. Xiao, N. Pinotsis, M. Wilmanns, F. Grater, and M. Rief (2011)
PNAS 108, 14139-14144
   Abstract »    Full Text »    PDF »
Ligand-modulated Parallel Mechanical Unfolding Pathways of Maltose-binding Proteins.
V. Aggarwal, S. R. Kulothungan, M. M. Balamurali, S. R. Saranya, R. Varadarajan, and S. R. K. Ainavarapu (2011)
J. Biol. Chem. 286, 28056-28065
   Abstract »    Full Text »    PDF »
Single-molecule Force Spectroscopy Reveals the Individual Mechanical Unfolding Pathways of a Surface Layer Protein.
C. Horejs, R. Ristl, R. Tscheliessnig, U. B. Sleytr, and D. Pum (2011)
J. Biol. Chem. 286, 27416-27424
   Abstract »    Full Text »    PDF »
Nanomechanics of the Cadherin Ectodomain: "CANALIZATION" BY Ca2+ BINDING RESULTS IN A NEW MECHANICAL ELEMENT.
J. Oroz, A. Valbuena, A. M. Vera, J. Mendieta, P. Gomez-Puertas, and M. Carrion-Vazquez (2011)
J. Biol. Chem. 286, 9405-9418
   Abstract »    Full Text »    PDF »
Deconvolution of dynamic mechanical networks.
M. Hinczewski, Y. von Hansen, and R. R. Netz (2010)
PNAS 107, 21493-21498
   Abstract »    Full Text »    PDF »
Full Reconstruction of a Vectorial Protein Folding Pathway by Atomic Force Microscopy and Molecular Dynamics Simulations.
W. Lee, X. Zeng, H.-X. Zhou, V. Bennett, W. Yang, and P. E. Marszalek (2010)
J. Biol. Chem. 285, 38167-38172
   Abstract »    Full Text »    PDF »
Sampling Protein Form and Function with the Atomic Force Microscope.
M. Baclayon, W. H. Roos, and G. J. L. Wuite (2010)
Mol. Cell. Proteomics 9, 1678-1688
   Abstract »    Full Text »    PDF »
Full distance-resolved folding energy landscape of one single protein molecule.
J. C. M. Gebhardt, T. Bornschlogl, and M. Rief (2010)
PNAS 107, 2013-2018
   Abstract »    Full Text »    PDF »
Single-molecule force spectroscopy distinguishes target binding modes of calmodulin.
J. P. Junker and M. Rief (2009)
PNAS 106, 14361-14366
   Abstract »    Full Text »    PDF »
Force Signaling in Biology.
J. Christof, M. Gebhardt, and M. Rief (2009)
Science 324, 1278-1280
   Abstract »    Full Text »    PDF »
Sending Signals Dynamically.
R. G. Smock and L. M. Gierasch (2009)
Science 324, 198-203
   Abstract »    Full Text »    PDF »
BIOCHEMISTRY: Unfolding the Secrets of Calmodulin.
R. B. Best and G. Hummer (2009)
Science 323, 593-594
   Abstract »    Full Text »    PDF »

To Advertise     Find Products

Science Signaling. ISSN 1937-9145 (online), 1945-0877 (print). Pre-2008: Science's STKE. ISSN 1525-8882