Neck-linker length dependence of processive Kinesin-5 motility.
ABSTRACT Processive motility of individual molecules is essential for the function of many kinesin motors. Processivity for kinesins relies on communication between the two heads of a dimeric molecule, such that binding strictly alternates. The main communicating elements are believed to be the two neck linkers connecting the motors' stalks and heads. A proposed mechanism for coordination is the transmission of stress through the neck linkers. It is believed that the efficiency of gating depends on the length of the neck linker. Recent studies have presented support for a simple model in which the length of the neck linker directly controls the degree of processivity. Based on a previously published Kinesin-1/Kinesin-5 chimera, Eg5Kin, we have analyzed the motility of 12 motor constructs: we have varied the length of the neck linker in the range between 9 and 21 amino acids using the corresponding native Kinesin-5 sequence (Xenopus laevis Eg5). We found, surprisingly, that neither velocity nor force generation depended on neck-linker length. We also found that constructs with short neck linkers, down to 12 amino acids, were still highly processive, while processivity was lost at a length of 9 amino acids. Run lengths were maximal with neck linkers close to the native Kinesin-5 length and decreased beyond that length. This finding generally confirms the coordinating role of the neck linker for kinesin motility but challenges the simplest model postulating a motor-type-independent optimal length. Instead, our results suggest that different kinesins might be optimized for different neck-linker lengths.
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ABSTRACT: The kinesin-3 family is one of the largest among the kinesin superfamily and its members play important roles in a wide range of cellular transport activities, yet the molecular mechanisms of kinesin-3 regulation and cargo transport are largely unknown. We performed a comprehensive analysis of mammalian kinesin-3 motors from three different subfamilies (KIF1, KIF13, and KIF16). Using Forster resonance energy transfer microscopy in live cells, we show for the first time to our knowledge that KIF16B motors undergo cargo-mediated dimerization. The molecular mechanisms that regulate the monomer-to-dimer transition center around the neck coil (NC) segment and its ability to undergo intramolecular interactions in the monomer state versus intermolecular interactions in the dimer state. Regulation of NC dimerization is unique to the kinesin-3 family and in the case of KIF13A and KIF13B requires the release of a proline-induced kink between the NC and subsequent coiled-coil 1 segments. We show that dimerization of kinesin-3 motors results in superprocessive motion, with average run lengths of ∼10 μm, and that this property is intrinsic to the dimeric kinesin-3 motor domain. This finding opens up studies on the mechanistic basis of motor processivity. Such high processivity has not been observed for any other motor protein and suggests that kinesin-3 motors are evolutionarily adapted to serve as the marathon runners of the cellular world.Proceedings of the National Academy of Sciences 04/2014; · 9.81 Impact Factor
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ABSTRACT: Kinesin motor proteins comprise an ATPase superfamily that goes hand in hand with microtubules in every eukaryote. The mitotic kinesins, by virtue of their potential therapeutic role in cancerous cells, have been a major focus of research for the past 28years since the discovery of the canonical Kinesin-1 heavy chain. Perhaps the simplest player in mitotic spindle assembly, Kinesin-5 (also known as Kif11, Eg5, or kinesin spindle protein, KSP) is a plus-end-directed motor localized to interpolar spindle microtubules and to the spindle poles. Comprised of a homotetramer complex, its function primarily is to slide anti-parallel microtubules apart from one another. Based on a multi-faceted analysis of this motor from numerous laboratories over the years, we have learned a great deal about the function of this motor at the atomic level for catalysis and as an integrated element of the cytoskeleton. These data have, in turn, informed the function of motile kinesins on the whole, as well as spearheaded integrative models of the mitotic apparatus in particular and regulation of the microtubule cytoskeleton in general. We review what is known about how this nanomotor works, its place inside the cytoskeleton of cells, and its small-molecule inhibitors that provide a toolbox for understanding motor function and for anticancer treatment in the clinic.Gene 08/2013; · 2.20 Impact Factor
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ABSTRACT: Mitotic cell division is the most fundamental task of all living cells. Cells have intricate and tightly regulated machinery to ensure that mitosis occurs with appropriate frequency and high fidelity. A core element of this machinery is the kinesin-5 motor protein, which plays essential roles in spindle formation and maintenance. In this review, we discuss how the structural and mechanical properties of kinesin-5 motors uniquely suit them to their mitotic role. We describe some of the small molecule inhibitors and regulatory proteins that act on kinesin-5, and discuss how these regulators may influence the process of cell division. Finally, we touch on some more recently described functions of kinesin-5 motors in non-dividing cells. Throughout, we highlight a number of open questions that impede our understanding of both this motor's function and the potential utility of kinesin-5 inhibitors. This article is protected by copyright. All rights reserved.Biology of the Cell 10/2013; · 3.49 Impact Factor