title: Kinesin-2 KIF3AB exhibits novel ATPase characteristics.
abstract: KIF3AB is an N-terminal processive kinesin-2 family member best known for its role in intraflagellar transport. There has been significant interest in KIF3AB in defining the key principles that underlie the processivity of KIF3AB in comparison with homodimeric processive kinesins. To define the ATPase mechanism and coordination of KIF3A and KIF3B stepping, a presteady-state kinetic analysis was pursued. For these studies, a truncated murine KIF3AB was generated. The results presented show that microtubule association was fast at 5.7 μm(-1) s(-1), followed by rate-limiting ADP release at 12.8 s(-1). ATP binding at 7.5 μm(-1) s(-1) was followed by an ATP-promoted isomerization at 84 s(-1) to form the intermediate poised for ATP hydrolysis, which then occurred at 33 s(-1). ATP hydrolysis was required for dissociation of the microtubule·KIF3AB complex, which was observed at 22 s(-1). The dissociation step showed an apparent affinity for ATP that was very weak (K½,ATP at 133 μm). Moreover, the linear fit of the initial ATP concentration dependence of the dissociation kinetics revealed an apparent second-order rate constant at 0.09 μm(-1) s(-1), which is inconsistent with fast ATP binding at 7.5 μm(-1) s(-1) and a Kd ,ATP at 6.1 μm. These results suggest that ATP binding per se cannot account for the apparent weak K½,ATP at 133 μm. The steady-state ATPase Km ,ATP, as well as the dissociation kinetics, reveal an unusual property of KIF3AB that is not yet well understood and also suggests that the mechanochemistry of KIF3AB is tuned somewhat differently from homodimeric processive kinesins.
authors.affiliation: From the Department of Biological Sciences and the Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York 12180 and.
authors.forename: Clayton D
authors.initials: CD
authors.lastname: Albracht
authors.affiliation: the Department of Biochemistry, University of Wisconsin, Madison, Wisconsin 53706.
authors.forename: Katherine C
authors.initials: KC
authors.lastname: Rank
authors.affiliation: From the Department of Biological Sciences and the Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York 12180 and.
authors.forename: Steven
authors.initials: S
authors.lastname: Obrzut
authors.affiliation: the Department of Biochemistry, University of Wisconsin, Madison, Wisconsin 53706 ivan_rayment@biochem.wisc.edu.
authors.forename: Ivan
authors.initials: I
authors.lastname: Rayment
authors.affiliation: From the Department of Biological Sciences and the Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York 12180 and sgilbert@rpi.edu.
authors.forename: Susan P
authors.initials: SP
authors.lastname: Gilbert
mesh_terms: D000244:Adenosine Diphosphate / Q000378:metabolism; D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D019281:Dimerization; D016547:Kinesins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D051379:Mice; D008870:Microtubules / Q000378:metabolism
chemical_list: C490202:Kif3a protein, mouse; C490672:Kif3b protein, mouse; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D016547:Kinesins
keywords: ATPase; Intracellular Trafficking; Intraflagellar Transport; Kinesin; Microtubule; Presteady-state Kinetics; Processivity