Mechanoenzymatics of titin kinase

Elias M. Puchner, Alexander Alexandrovich, Lin Kho Ay, Ulf Hensen, Lars V. Schäfer, Birgit Brandmeier, Frauke Gräter, Helmut Grubmüller, Hermann E. Gaub, Mathias Gautel

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240 Scopus citations

Abstract

Biological responses to mechanical stress require strain-sensing molecules, whose mechanically induced conformational changes are relayed to signaling cascades mediating changes in cell and tissue properties. In vertebrate muscle, the giant elastic protein titin is involved in strain sensing via its C-terminal kinase domain (TK) at the sarcomeric M-band and contributes to the adaptation of muscle in response to changes in mechanical strain. TK is regulated in a unique dual autoinhibition mechanism by a C-terminal regulatory tail, blocking the ATP binding site, and tyrosine autoinhibition of the catalytic base. For access to the ATP binding site and phosphorylation of the autoinhibitory tyrosine, the C-terminal autoinhibitory tail needs to be removed. Here, we use AFM-based single-molecule force spectroscopy, molecular dynamics simulations, and enzymatics to study the conformational changes during strain-induced activation of human TK. We show that mechanical strain activates ATP binding before unfolding of the structural titin domains, and that TK can thus act as a biological force sensor. Furthermore, we identify the steps in which the autoinhibition of TK is mechanically relieved at low forces, leading to binding of the cosubstrate ATP and priming the enzyme for subsequent autophosphorylation and substrate turnover.

Original languageEnglish (US)
Pages (from-to)13385-13390
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume105
Issue number36
DOIs
StatePublished - Sep 9 2008

Keywords

  • Atomic force microscopy
  • Force-probe molecular dynamics simulation
  • Muscle signaling
  • Protein kinase regulation
  • Single-molecular force spectroscopy

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    Puchner, E. M., Alexandrovich, A., Ay, L. K., Hensen, U., Schäfer, L. V., Brandmeier, B., Gräter, F., Grubmüller, H., Gaub, H. E., & Gautel, M. (2008). Mechanoenzymatics of titin kinase. Proceedings of the National Academy of Sciences of the United States of America, 105(36), 13385-13390. https://doi.org/10.1073/pnas.0805034105