In this study, we aimed to study the role of inorganic phosphate (Pi) in the production of oscillatory work and cross-bridge (CB) kinetics of striated muscle. We applied small-amplitude sinusoidal length oscillations to rabbit psoas single myofibrils and muscle fibers, and the resulting force responses were analyzed during maximal Ca2+ activation (pCa 4.65) at 15°C. Three exponential processes, A, B, and C, were identified from the tension transients, which were studied as functions of Pi concentration ([Pi]). In myofibrils, we found that process C, corresponding to phase 2 of step analysis during isometric contraction, is almost a perfect single exponential function compared with skinned fibers, which exhibit distributed rate constants, as described previously. The [Pi] dependence of the apparent rate constants 2πb and 2πc, and that of isometric tension, was studied to characterize the force generation and Pi release steps in the CB cycle, as well as the inhibitory effect of Pi. In contrast to skinned fibers, Pi does not accumulate in the core of myofibrils, allowing sinusoidal analysis to be performed nearly at [Pi] = 0. Process B disappeared as [Pi] approached 0 mM in myofibrils, indicating the significance of the role of Pi rebinding to CBs in the production of oscillatory work (process B). Our results also suggest that Pi competitively inhibits ATP binding to CBs, with an inhibitory dissociation constant of ∼2.6 mM. Finally, we found that the sinusoidal waveform of tension is mostly distorted by second harmonics and that this distortion is closely correlated with production of oscillatory work, indicating that the mechanism of generating force is intrinsically nonlinear. A nonlinear force generation mechanism suggests that the length-dependent intrinsic rate constant is asymmetric upon stretch and release and that there may be a ratchet mechanism involved in the CB cycle.
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1 March 2021
Article|
February 12 2021
Phosphate has dual roles in cross-bridge kinetics in rabbit psoas single myofibrils
Masataka Kawai
,
1
Department of Anatomy and Cell Biology, University of Iowa, Iowa City, IA
Correspondence to Masataka Kawai: masataka-kawai@uiowa.edu
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Robert Stehle
,
Robert Stehle
2
Institute of Vegetative Physiology, University of Köln, Köln, Germany
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Gabriele Pfitzer
,
Gabriele Pfitzer
2
Institute of Vegetative Physiology, University of Köln, Köln, Germany
3
Institute of Neurophysiology, University of Köln, Köln, Germany
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Bogdan Iorga
Bogdan Iorga
2
Institute of Vegetative Physiology, University of Köln, Köln, Germany
4
Department of Molecular and Cell Physiology, Hannover Medical School, Hannover, Germany
5
Department of Physical Chemistry, Faculty of Chemistry, University of Bucharest, Bucharest, Romania
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Masataka Kawai
1
Department of Anatomy and Cell Biology, University of Iowa, Iowa City, IA
Robert Stehle
2
Institute of Vegetative Physiology, University of Köln, Köln, Germany
Gabriele Pfitzer
2
Institute of Vegetative Physiology, University of Köln, Köln, Germany
3
Institute of Neurophysiology, University of Köln, Köln, Germany
Bogdan Iorga
2
Institute of Vegetative Physiology, University of Köln, Köln, Germany
4
Department of Molecular and Cell Physiology, Hannover Medical School, Hannover, Germany
5
Department of Physical Chemistry, Faculty of Chemistry, University of Bucharest, Bucharest, Romania
Correspondence to Masataka Kawai: masataka-kawai@uiowa.edu
This work is part of a special collection on myofilament function and disease.
Received:
September 05 2020
Revision Received:
December 04 2020
Accepted:
January 15 2021
Online Issn: 1540-7748
Print Issn: 0022-1295
Funding:
National Institutes of Health
(HL70041)
© 2021 Kawai et al.
2021
This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms/). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 International license, as described at https://creativecommons.org/licenses/by-nc-sa/4.0/).
J Gen Physiol (2021) 153 (3): e202012755.
Article history
Received:
September 05 2020
Revision Received:
December 04 2020
Accepted:
January 15 2021
Citation
Masataka Kawai, Robert Stehle, Gabriele Pfitzer, Bogdan Iorga; Phosphate has dual roles in cross-bridge kinetics in rabbit psoas single myofibrils. J Gen Physiol 1 March 2021; 153 (3): e202012755. doi: https://doi.org/10.1085/jgp.202012755
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