Molecular mechanics of two smooth muscle heavy meromyosin constructs that differ by an insert in the motor domain.
Phasic and tonic smooth muscles express two myosin heavy chain isoforms that differ by only a seven amino acid insert in a flexible surface loop located near the nucleotide-binding site. The inserted isoform found predominantly in phasic muscle has two times the actin-activated ATPase activity and in vitro actin filament velocity as the non-insert isoform found mainly in tonic muscle (Kelley, C.A., Takahashi, M., Yu, J.H. & Adelstein, R.S. 1993. J Biol Chem 268, 12848, Rovner, A.S., Freyzon, Y. & Trybus, K.M. 1997. J Musc Res Cell Motil 18, 103). We used a laser trap to characterize the molecular mechanics of the inserted isoform [(+)insert] and of a mutant lacking the insert [(-)insert], which is analogous to the isoform found in tonic muscles. The constructs were expressed in the baculovirus/insect cell system. Unitary displacements (Duni) were similar for both the constructs (approximately 10 nm) but the attachment time (ton) for the (-)insert was two times that of the (+)insert. These data suggest that the insert in the nucleotide-binding loop does not affect the inherent mechanics of the myosin molecule but rather the kinetics of the cross-bridge cycle.
Lauzon
A M
AM
Department of Molecular Physiology and Biophysics, University of Vermont, Burlington 05405, USA.
Trybus
K M
KM
Warshaw
D M
DM
D000199:Actins / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002021:Buffers; D000070559:Gizzard, Non-avian / Q000378:metabolism; D066298:In Vitro Techniques; D007536:Isomerism; D007700:Kinetics; D009119:Muscle Contraction / Q000502:physiology; D009130:Muscle, Smooth / Q000378:metabolism* / Q000648:ultrastructure; D015879:Myosin Subfragments / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D009218:Myosins / Q000378:metabolism
D000199:Actins; D002021:Buffers; D015879:Myosin Subfragments; D000255:Adenosine Triphosphate; D009218:Myosins