Nanospring behaviour of ankyrin repeats

Ankyrin repeats are an amino-acid motif believed to function in protein recognition; they are present in tandem copies in diverse proteins in nearly all phyla. Ankyrin repeats contain antiparallel α-helices that can stack to form a superhelical spiral. Visual inspection of the extrapolated structure of 24 ankyrin-R repeats indicates the possibility of spring-like behaviour of the putative superhelix. Moreover, stacks of 17–29 ankyrin repeats in the cytoplasmic domains of transient receptor potential (TRP) channels have been identified as candidates for a spring that gates mechanoreceptors in hair cells as well as in Drosophila bristles. Here we report that tandem ankyrin repeats exhibit tertiary-structure-based elasticity and behave as a linear and fully reversible spring in single-molecule measurements by atomic force microscopy. We also observe an unexpected ability of unfolded repeats to generate force during refolding, and report the first direct measurement of the refolding force of a protein domain. Thus, we show that one of the most common amino-acid motifs has spring properties that could be important in mechanotransduction and in the design of nanodevices.

[1]  A. Baines,et al.  Spectrin and ankyrin-based pathways: metazoan inventions for integrating cells into tissues. , 2001, Physiological reviews.

[2]  K. Schulten,et al.  In search of the hair-cell gating spring elastic properties of ankyrin and cadherin repeats. , 2005, Structure.

[3]  E. Siggia,et al.  Entropic elasticity of lambda-phage DNA. , 1994, Science.

[4]  J. Ashmore.  Hearing: Channel at the hair's end , 2004, Nature.

[5]  Jonathon Howard,et al.  Hypothesis: A helix of ankyrin repeats of the NOMPC-TRP ion channel is the gating spring of mechanoreceptors , 2004, Current Biology.

[6]  Heidi L. Rehm,et al.  TRPA1 is a candidate for the mechanosensitive transduction channel of vertebrate hair cells , 2004, Nature.

[7]  Carlos Bustamante,et al.  Direct Observation of the Three-State Folding of a Single Protein Molecule , 2005, Science.

[8]  D. Corey,et al.  Hearing: Tightrope act , 2004, Nature.

[9]  Piotr E. Marszalek,et al.  Stretching single molecules into novel conformations using the atomic force microscope , 2000, Nature Structural Biology.

[10]  Matthias Rief,et al.  The myosin coiled-coil is a truly elastic protein structure , 2002, Nature materials.

[11]  Laura S Itzhaki,et al.  Sequential unfolding of ankyrin repeats in tumor suppressor p16. , 2003, Structure.

[12]  Mariano Carrion-Vazquez,et al.  The mechanical hierarchies of fibronectin observed with single-molecule AFM. , 2002, Journal of molecular biology.

[13]  Doug Barrick,et al.  An experimentally determined protein folding energy landscape. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[14]  M. Rief,et al.  Reversible unfolding of individual titin immunoglobulin domains by AFM. , 1997, Science.

[15]  Julio M Fernandez,et al.  Force-Clamp Spectroscopy Monitors the Folding Trajectory of a Single Protein , 2004, Science.

[16]  Peter Michaely,et al.  Crystal structure of a 12 ANK repeat stack from human ankyrinR , 2002, The EMBO journal.

[17]  S. Radford,et al.  Force mode atomic force microscopy as a tool for protein folding studies , 2003 .

[18]  M. Rief,et al.  Single molecule force spectroscopy of spectrin repeats: low unfolding forces in helix bundles. , 1999, Journal of molecular biology.

[19]  Wolfgang A. Linke,et al.  Reverse engineering of the giant muscle protein titin , 2002, Nature.

[20]  A. J. Hudspeth,et al.  Compliance of the hair bundle associated with gating of mechanoelectrical transduction channels in the Bullfrog's saccular hair cell , 1988, Neuron.

[21]  H. Gaub,et al.  A metal-chelating microscopy tip as a new toolbox for single-molecule experiments by atomic force microscopy. , 2000, Biophysical journal.

[22]  Andres F. Oberhauser,et al.  The molecular elasticity of the extracellular matrix protein tenascin , 1998, Nature.

[23]  Matthias Rief,et al.  Sensing specific molecular interactions with the atomic force microscope , 1995 .

[24]  Jane Clarke,et al.  Hidden complexity in the mechanical properties of titin , 2003, Nature.

[25]  Klaus Schulten,et al.  Mechanical unfolding intermediates in titin modules , 1999, Nature.

[26]  Daniel C. Desrosiers,et al.  The ankyrin repeat as molecular architecture for protein recognition , 2004, Protein science : a publication of the Protein Society.

[27]  Z. Dienes,et al.  Reversible oriented surface immobilization of functional proteins on oxide surfaces. , 1997, Analytical chemistry.

[28]  M. Rief,et al.  The folding pathway of a fast‐folding immunoglobulin domain revealed by single‐molecule mechanical experiments , 2005, EMBO reports.