Carrington, G., Fatima, U., Caramujo, I. et al. (3 more authors) (2024) A multiscale approach reveals the molecular architecture of the autoinhibited kinesin KIF5A. Journal of Biological Chemistry, 300 (3). 105713. ISSN 0021-9258
Abstract
Kinesin-1 is a microtubule motor that transports cellular cargo along microtubules. KIF5A is one of three kinesin-1 isoforms in humans, all of which are autoinhibited by an interaction between the motor and an IAK motif in the proximal region of the C-terminal tail. The C-terminal tail of KIF5A is ∼80 residues longer than the other two kinesin-1 isoforms (KIF5B and KIF5C) and it is unclear if it contributes to autoinhibition. Mutations in KIF5A cause neuronal diseases and could affect autoinhibition, as reported for a mutation that skips exon 27, altering its C-terminal sequence. Here, we combined negative-stain electron microscopy, crosslinking mass spectrometry (XL-MS) and AlphaFold2 structure prediction to determine the molecular architecture of the full-length autoinhibited KIF5A homodimer, in the absence of light chains. We show that KIF5A forms a compact, bent conformation, through a bend between coiled-coils 2 and 3, around P687. XL-MS of WT KIF5A revealed extensive interactions between residues in the motor, between coiled-coil 1 and the motor, between coiled-coils 1 and 2, with coiled-coils 3 and 4, and the proximal region of the C-terminal tail and the motor in the autoinhibited state, but not between the distal C-terminal region and the rest of the molecule. While negative-stain electron microscopy of exon-27 KIF5A splice mutant showed the presence of autoinhibited molecules, XL-MS analysis suggested that its autoinhibited state is more labile. Our model offers a conceptual framework for understanding how mutations within the motor and stalk domain may affect motor activity.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2024 THE AUTHORS. Published by Elsevier Inc on behalf of American Society for Biochemistry and Molecular Biology. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/). |
Keywords: | Kinesin; KIF5A; negative-stain electron microscopy; crosslinking mass spectrometry; protein crosslinking; Alphafold; structure-function; cytoskeleton; inhibition mechanism |
Dates: |
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Institution: | The University of Leeds |
Academic Units: | The University of Leeds > Faculty of Biological Sciences (Leeds) > School of Molecular and Cellular Biology (Leeds) > Cell Biology (Leeds) |
Funding Information: | Funder Grant number Wellcome Trust 223125/Z/21/Z Wellcome Trust 104918/Z/14/Z Wellcome Trust 223810/Z/21/Z |
Depositing User: | Symplectic Publications |
Date Deposited: | 02 Feb 2024 12:50 |
Last Modified: | 28 Feb 2024 13:56 |
Status: | Published |
Publisher: | Elsevier |
Identification Number: | 10.1016/j.jbc.2024.105713 |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:208602 |