Single-molecule measurement of the stiffness of the rigor myosin head.
The force-extension curve of single myosin subfragment-1 molecules, interacting in the rigor state with an actin filament, has been investigated at low [ATP] by applying a slow triangle-wave movement to the optical traps holding a bead-actin-bead dumbbell. In combination with a measurement of the overall stiffness of the dumbbell, this allowed characterization of the three extensible elements, the actin-bead links and the myosin. Simultaneously, another method, based on an analysis of bead position covariance, gave satisfactory agreement. The mean covariance-based estimate for the myosin stiffness was 1.79 pN/nm (SD = 0.7 pN/nm; SE = 0.06 pN/nm (n = 166 myosin molecules)), consistent with a recent report (1.7 pN/nm) from rabbit muscle fibers. In the triangle-wave protocol, the motion of the trapped beads during interactions was linear within experimental error over the physiological range of force applied to myosin (+/-10 pN), consistent with a Hookean model; any nonlinear terms could not be characterized. Bound states subjected to forces that resisted the working stroke (i.e., positive forces) detached at a significantly lower force than when subjected to negative forces, which is indicative of a strain-dependent dissociation rate.
Randall Division, King's College London, London, United Kingdom. john.sleep@kcl.ac.uk
Alexandre
A
Lewalle
Walter
W
Steffen
Olivia
O
Stevenson
Zhenqian
Z
Ouyang
John
J
Sleep
D008841:Actin Cytoskeleton / Q000378:metabolism; D000199:Actins / Q000737:chemistry; D000205:Actomyosin / Q000737:chemistry; D000255:Adenosine Triphosphate / Q000737:chemistry; D000818:Animals; D001703:Biophysics / Q000379:methods*; D009119:Muscle Contraction; D018485:Muscle Fibers, Skeletal / Q000378:metabolism; D009127:Muscle Rigidity; D009132:Muscles / Q000378:metabolism; D015879:Myosin Subfragments / Q000737:chemistry; D009218:Myosins / Q000737:chemistry*; D011487:Protein Conformation; D011817:Rabbits; D013816:Thermodynamics
D000199:Actins; D015879:Myosin Subfragments; D000255:Adenosine Triphosphate; D000205:Actomyosin; D009218:Myosins