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Updated: May 28, 2026

A Protocol for Using Förster Resonance Energy Transfer (FRET)-force Biosensors to Measure Mechanical Forces across the Nuclear LINC Complex
Published on: April 11, 2017
A conditional gating mechanism assures the integrity of the molecular force-sensor titin kinase
Stefan W Stahl1, Elias M Puchner, Alexander Alexandrovich
1Center for NanoScience, Ludwig-Maximilians-University Munich, Munich, Germany.
Abstract:
As more and more recent investigations point out, force plays an important role in cellular regulation mechanisms. Biological responses to mechanical stress are often based on force-induced conformational changes of single molecules. The force sensor, titin kinase, is involved in a signaling complex that regulates protein turnover and transcriptional adaptation in striated muscle. The structural architecture of such a force sensor determines its response to force and must assure both activity and mechanical integrity, which are prerequisites for its function. Here, we use single-molecule force-clamp spectroscopy to show that titin kinase is organized in such a way that the regulatory domains have to unfold before secondary structure elements that determine the overall fold and catalytic function. The stepwise unfolding over many barriers with a topologically determined sequence assures that the protein can react to force by conformational changes while maintaining its structural integrity.
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