Contribution of the myosin VI tail domain to processive stepping and intramolecular tension sensing.
Myosin VI is proposed to act as both a molecular transporter and as an anchor in vivo. A portion of the molecule C-terminal to the canonical lever arm, termed the medial tail (MT), has been proposed to act as either a lever arm extension or as a dimerization motif. We describe constructs in which the MT is interrupted by a glycine-rich molecular swivel. Disruption of the MT results in decreased processive run lengths measured using single-molecule fluorescence microscopy and a decreased step size under applied load as measured in an optical trap. We used single-molecule gold nanoparticle tracking and optical trapping to examine the mechanism of coordination between the heads of dimeric myosin VI. We detect two rate-limiting kinetic processes at low (< 200 micromolar) ATP concentrations. Our data can be explained by a model in which intramolecular tension greatly increases the affinity of the lead head for ADP, likely by slowing ADP release from the lead head. This mechanism likely increases both the motor's processivity and its ability to act as an anchor under physiological conditions.
Dunn
Alexander R
AR
Department of Biochemistry, Stanford University School of Medicine, Stanford, CA 94305, USA.
Chuan
Peiying
P
Bryant
Zev
Z
Spudich
James A
JA
D000255:Adenosine Triphosphate / Q000378:metabolism; D001696:Biomechanical Phenomena; D003198:Computer Simulation; D019281:Dimerization; D007700:Kinetics; D008856:Microscopy, Fluorescence; D008956:Models, Chemical; D009010:Monte Carlo Method; D018995:Myosin Heavy Chains / Q000737:chemistry*; D052898:Optical Tweezers; D017434:Protein Structure, Tertiary
C089785:myosin VI; D000255:Adenosine Triphosphate; D018995:Myosin Heavy Chains
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