Temperature change does not affect force between regulated actin filaments and heavy meromyosin in single-molecule experiments

Masataka Kawai; Takanori Kido; Martin Vogel; Rainer H A Fink; Shin'ichi Ishiwata

doi:10.1113/jphysiol.2006.111708

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Temperature change does not affect force between regulated actin filaments and heavy meromyosin in single-molecule experiments

Journal article

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Temperature change does not affect force between regulated actin filaments and heavy meromyosin in single-molecule experiments

Masataka Kawai, Takanori Kido, Martin Vogel, Rainer H A Fink and Shin'ichi Ishiwata

The Journal of physiology, Vol.574(Pt 3), pp.877-887

08/01/2006

DOI: 10.1113/jphysiol.2006.111708

PMCID: PMC1817734

PMID: 16709631

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Abstract

The temperature dependence of sliding velocity, force and the number of cross-bridges was studied on regulated actin filaments (reconstituted thin filaments) when they were placed on heavy meromyosin (HMM) attached to a glass surface. The regulated actin filaments were used because our previous study on muscle fibres demonstrated that the temperature effect was much reduced in the absence of regulatory proteins. A fluorescently labelled thin filament was attached to the gelsolin-coated surface of a polystyrene bead. The bead was trapped by optical tweezers, and HMM-thin filament interaction was performed at 20-35 degrees C to study the temperature dependence of force at the single-molecule level. Our experiments showed that there was a small increase in force with temperature (Q10 = 1.43) and sliding velocity (Q10 = 1.46). The small increase in force was correlated with the small increase in the number of cross-bridges (Q10 = 1.49), and when force was divided by the number of cross-bridges, the result did not depend on the temperature (Q(10) = 1.03). These results demonstrate that the force each cross-bridge generates is fixed and independent of temperature. Our additional experiments demonstrate that tropomyosin (Tm) in the presence of troponin (Tn) and Ca2+ enhances both force and velocity, and a truncated mutant, Delta23Tm, diminishes force and velocity. These results are consistent with the hypothesis that Tm in the presence of Tn and Ca2+ exerts a positive allosteric effect on actin to make actomyosin linkage more secure so that larger forces can be generated.

Temperature

Muscle, Skeletal - chemistry

Actin Cytoskeleton - chemistry

Protein Binding

Stress, Mechanical

Myosin Subfragments - chemistry

Sarcomeres - chemistry

Molecular Motor Proteins - chemistry

Molecular Biology - methods

Details

Title: Subtitle: Temperature change does not affect force between regulated actin filaments and heavy meromyosin in single-molecule experiments
Creators: Masataka Kawai - Department of Anatomy and Cell Biology, University of Iowa, Iowa City, IA 52242, USA. masataka-kawai@uiowa.edu
Takanori Kido
Martin Vogel
Rainer H A Fink
Shin'ichi Ishiwata
Resource Type: Journal article
Publication Details: The Journal of physiology, Vol.574(Pt 3), pp.877-887
DOI: 10.1113/jphysiol.2006.111708
PMID: 16709631
PMCID: PMC1817734
NLM abbreviation: J Physiol
ISSN: 0022-3751
eISSN: 1469-7793
Publisher: England
Grant note: R01 HL070041 / NHLBI NIH HHS R01 HL070041-04 / NHLBI NIH HHS HL70041 / NHLBI NIH HHS
Language: English
Date published: 08/01/2006
Academic Unit: Anatomy and Cell Biology; Internal Medicine
Record Identifier: 9984025560702771

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