Orientation changes of fluorescent probes at five sites on the myosin regulatory light chain during contraction of single skeletal muscle fibres

Journal of Molecular Biology
C Sabido-DavidM Irving

Abstract

Changes in the orientation of the myosin regulatory light chain (RLC) in single muscle fibres were measured using polarised fluorescence from acetamidotetramethylrhodamine (ATR). Mutants of chicken skeletal RLC containing single cysteine residues at positions 2, 73, 94, 126 and 155 were labelled with either the 5 or 6-isomer of iodo-ATR, giving ten different probes. The labelled RLCs were exchanged into demembranated fibres from rabbit psoas muscle without significant effect on active force generation. Fluorescence polarisation measurements showed that nine out of the ten probe dipoles were more perpendicular to the fibre axis in the absence of ATP (in rigor) than in either relaxation or active contraction. The orientational distribution of the RLC region of the myosin head in active contraction is closer to the relaxed than to the rigor orientation, and is not equivalent to a linear combination of the relaxed and rigor orientations. Rapid length steps were applied to the fibres to synchronise the motions of myosin heads attached to actin. In active contraction the fluorescence polarisation changed both during the step, indicating elastic distortion of the RLC region of the myosin head, and during the subsequent rapid force rec...Continue Reading

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Citations

Jan 19, 2010·Proceedings of the National Academy of Sciences of the United States of America·Joseph M LaaksoE Michael Ostap
Jun 29, 2000·Annual Review of Biochemistry·M A Geeves, K C Holmes
May 7, 2009·Annual Review of Biophysics·David D ThomasVicci L Korman
Nov 6, 2002·Proceedings of the National Academy of Sciences of the United States of America·Diane S Lidke, David D Thomas
May 23, 2007·Proceedings of the National Academy of Sciences of the United States of America·Barbara ColombiniGiovanni Cecchi
Dec 15, 2015·Frontiers in Cardiovascular Medicine·Janhavi NagwekarJulian Borejdo
Jan 8, 1999·Journal of Molecular Biology·C C AshleyG Rapp
Feb 16, 2005·Journal of Muscle Research and Cell Motility·A A ShepardJ Borejdo
Nov 29, 2013·American Journal of Physiology. Regulatory, Integrative and Comparative Physiology·Divya DuggalJulian Borejdo
Dec 7, 2000·Progress in Biophysics and Molecular Biology·J N ForkeyY E Goldman
Apr 30, 2003·Molecular Cell·Roisean E FergusonMalcolm Irving
Jul 29, 1999·Biophysical Journal·C Picart, D E Discher
Apr 2, 2003·Biophysical Journal·J Borejdo, I Akopova
Jun 21, 2011·Journal of Molecular and Cellular Cardiology·K MiddeJ Borejdo

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