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Modulating the Mechanical Stability of Extracellular Matrix Protein Tenascin-C in a Controlled and Reversible Fashion

TitleModulating the Mechanical Stability of Extracellular Matrix Protein Tenascin-C in a Controlled and Reversible Fashion
Publication TypeJournal Article
Year of Publication2009
AuthorsZhuang, SL, Peng, Q, Cao, Y, Li, HB
JournalJournal of Molecular Biology
Date PublishedJul
Type of ArticleArticle
ISBN Number0022-2836
KeywordsDYNAMICS, ELASTICITY, ENGINEERING PROTEINS, FNIII DOMAIN, FRAGMENTS, MECHANICAL STABILITY, mechanical unfolding, microscopy, MOLECULE FORCE SPECTROSCOPY, MUSCLE, rational design, recombination, SINGLE PROTEIN, single-molecule force spectroscopy, tenascin

Stretching force can induce conformational changes of proteins and is believed to be an important biological signal in the mechanotransduction network. Tenascin-C is a large extracellular matrix protein and is subject to stretching force under its physiological condition. Regulating the mechanical properties of the fibronectin type III domains of tenascin-C will alter its response to mechanical stretching force and thus may provide the possibility of regulating the biological activities of tenascin-C in living cells. However, tuning the mechanical stability of proteins in a rational and systematic fashion remains challenging. Using the third fibronectin type III domain (TNfn3) of tenascin-C as a model system, here we report a successful engineering of a mechanically stronger extracellular matrix protein via engineered metal chelation. Combining steered molecular dynamics simulations, protein engineering and single-molecule atomic force microscopy, we have rationally engineered a bihistidine-based metal chelation site into TNfn3. We used its metal chelation capability to selectively increase the unfolding energy barrier for the rate-limiting step during the mechanical unfolding of TNfn3. The resultant TNfn3 mutant exhibits enhanced mechanical stability. Using a stronger metal chelator, one can convert TNfn3 back to a state of lower mechanical stability. This is the first step toward engineering extracellular matrix proteins with defined mechanical properties, which can be modulated reversibly by external stimuli, and will provide the possibility of using external stimuli to regulate the biological functions of extracellular matrix proteins. (C) 2009 Elsevier Ltd. All rights reserved.

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