Human myosin Vc is a low duty ratio, nonprocessive molecular motor.
Myosin Vc is the product of one of the three genes of the class V myosin found in vertebrates. It is widely found in secretory and glandular tissues, with a possible involvement in transferrin trafficking. Transient and steady-state kinetic studies of human myosin Vc were performed using a truncated, single-headed construct. Steady-state actin-activated ATPase measurements revealed a V(max) of 1.8 +/- 0.3 s(-1) and a K(ATPase) of 43 +/- 11 microm. Unlike previously studied vertebrate myosin Vs, the rate-limiting step in the actomyosin Vc ATPase pathway is the release of inorganic phosphate (~1.5 s(-1)), rather than the ADP release step (~12.0-16.0 s(-1)). Nevertheless, the ADP affinity of actomyosin Vc (K(d) = 0.25 +/- 0.02 microm) reflects a higher ADP affinity than seen in other myosin V isoforms. Using the measured kinetic rates, the calculated duty ratio of myosin Vc was approximately 10%, indicating that myosin Vc spends the majority of the actomyosin ATPase cycle in weak actin-binding states, unlike the other vertebrate myosin V isoforms. Consistent with this, a fluorescently labeled double-headed heavy meromyosin form showed no processive movements along actin filaments in a single molecule assay, but it did move actin filaments at a velocity of approximately 24 nm/s in ensemble assays. Kinetic simulations reveal that the high ADP affinity of actomyosin Vc may lead to elevations of the duty ratio of myosin Vc to as high as 64% under possible physiological ADP concentrations. This, in turn, may possibly imply a regulatory mechanism that may be sensitive to moderate changes in ADP concentration.
Takagi
Yasuharu
Y
Laboratory of Molecular Physiology, NHLBI, NIH, Bethesda, MD 20892-8015, USA.
Yang
Yi
Y
Fujiwara
Ikuko
I
Jacobs
Damon
D
Cheney
Richard E
RE
Sellers
James R
JR
Kovács
Mihály
M
D000199:Actins / Q000378:metabolism; D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D004789:Enzyme Activation; D015870:Gene Expression; D006801:Humans; D006868:Hydrolysis; D007700:Kinetics; D009046:Motor Neurons / Q000378:metabolism*; D024701:Myosin Type V / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D010802:Phylogeny; D011485:Protein Binding
D000199:Actins; C459104:MYO5C protein, human; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D024701:Myosin Type V