Kinetic mechanism of human myosin IIIA

Andréa C. Dosé, Shobana Ananthanarayanan, Judy E. Moore, Beth Burnside, Christopher M. Yengo

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Abstract

Myosin IIIA is specifically expressed in photoreceptors and cochlea and is important for the phototransduction and hearing processes. In addition, myosin IIIA contains a unique N-terminal kinase domain and C-terminal tail actin-binding motif. We examined the kinetic properties of baculovirus expressed human myosin IIIA containing the kinase, motor, and two IQ domains. The maximum actin-activated ATPase rate is relatively slow (kcat = 0.77 ± 0.08 s-1), and high actin concentrations are required to fully activate the ATPase rate (KATPase = 34 ± 11 μM). However, actin co-sedimentation assays suggest that myosin III has a relatively high steady-state affinity for actin in the presence of ATP (Kactin ∼ 7 μM). The rate of ATP binding to the motor domain is quite slow both in the presence and absence of actin (K1 k+2 = 0.020 and 0.001 μM-1·s-1, respectively). The rate of actin-activated phosphate release is more than 100-fold faster (85 s -1) than the kcat, whereas ADP release in the presence of actin follows a two-step mechanism (7.0 and 0.6 s-1). Thus, our data suggest a transition between two actomyosin-ADP states is the rate-limiting step in the actomyosin III ATPase cycle. Our data also suggest the myosin III motor spends a large fraction of its cycle in an actomyosin ADP state that has an intermediate affinity for actin (Kd ∼ 5 μM). The long lived actomyosin-ADP state may be important for the ability of myosin III to function as a cellular transporter and actin cross-linker in the actin bundles of sensory cells.

Original languageEnglish (US)
Pages (from-to)216-231
Number of pages16
JournalJournal of Biological Chemistry
Volume282
Issue number1
DOIs
StatePublished - Jan 5 2007

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All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Dosé, A. C., Ananthanarayanan, S., Moore, J. E., Burnside, B., & Yengo, C. M. (2007). Kinetic mechanism of human myosin IIIA. Journal of Biological Chemistry, 282(1), 216-231. https://doi.org/10.1074/jbc.M605964200