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Single-Molecule Studies on the Motion and Force Generation of the Kinesin-3 Motor KIF1A

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Optical Tweezers

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2478))

Abstract

KIF1A is a neuron-specific member of the kinesin-3 family of microtubule (MT) plus-end-directed motor proteins. It powers the migration of nuclei in differentiating brain stem cells and the transport of synaptic precursors and dense core vesicles in axons. Its dysfunction causes severe neurodevelopmental and neurodegenerative diseases termed KIF1A-associated neurological disorders (KAND). KAND mutations span the entirety of the KIF1A protein sequence, of which the majority are located within the motor domain and are thus predicted to affect the motor’s motility and force-generating properties. Unfortunately, the molecular etiologies of KAND remain poorly understood, in part because KIF1A’s molecular mechanism remains unclear. Here, we describe detailed methods for how to express a tail-truncated dimeric KIF1A in E. coli cells and provide step-by-step protocols for performing single-molecule studies with total internal reflection fluorescence microscopy and optical tweezers assays, which, when combined with structure-function studies, help to decipher KIF1A’s molecular mechanism.

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Acknowledgments

L. Rao and A. Gennerich are supported by National Institutes of Health grants R01GM098469 and R01NS114636.

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Authors and Affiliations

  1. Department of Biochemistry and Gruss-Lipper Biophotonics Center, Albert Einstein College of Medicine, Bronx, NY, USA

    Lu Rao & Arne Gennerich

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  1. Lu Rao
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  2. Arne Gennerich
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Correspondence to Lu Rao or Arne Gennerich .

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  1. Department of Biochemistry, Albert Einstein College of Medicine, Bronx, NY, USA

    Arne Gennerich

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Rao, L., Gennerich, A. (2022). Single-Molecule Studies on the Motion and Force Generation of the Kinesin-3 Motor KIF1A. In: Gennerich, A. (eds) Optical Tweezers. Methods in Molecular Biology, vol 2478. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2229-2_21

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  • DOI: https://doi.org/10.1007/978-1-0716-2229-2_21

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2228-5

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