title: Kinetic characterization of tail swing steps in the ATPase cycle of Dictyostelium cytoplasmic dynein.
abstract: According to the power stroke model of dynein deduced from electron microscopic and fluorescence resonance energy transfer studies, the power stroke and the recovery stroke are expected to take place at the two isomerization steps of the ATPase cycle at the primary ATPase site. Here, we have conducted presteady-state kinetic analyses of these two isomerization steps with the single-headed motor domain of Dictyostelium cytoplasmic dynein by employing fluorescence resonance energy transfer to probe ATPase steps at the primary site and tail positions. Our results show that the recovery stroke at the first isomerization step proceeds quickly ( approximately 180 s(-1)), whereas the power stroke at the second isomerization step is very slow ( approximately 0.2 s(-1)) in the absence of microtubules, and that the presence of microtubules accelerates the second but not the first step. Moreover, a comparison of the microtubule-induced acceleration of the power stroke step and that of steady-state ATP hydrolysis implies the intriguing possibility that microtubules simultaneously accelerate the ATPase activity not only at the primary site but also at other site(s) in the motor domain.
authors.affiliation: Department of Life Sciences, Graduate School of Arts and Sciences, University of Tokyo, Komaba 3-8-1, Tokyo, Japan.
authors.forename: Toshifumi
authors.initials: T
authors.lastname: Mogami
authors.forename: Takahide
authors.initials: T
authors.lastname: Kon
authors.forename: Kohji
authors.initials: K
authors.lastname: Ito
authors.forename: Kazuo
authors.initials: K
authors.lastname: Sutoh
mesh_terms: D000818:Animals; D004023:Dictyostelium / Q000201:enzymology*; D004398:Dyneins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D010957:Plasmids; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D013050:Spectrometry, Fluorescence
chemical_list: D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; D004398:Dyneins