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Nucleotide-Dependent Dimerization and Conformational Switching of Atlastin

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Dynamin Superfamily GTPases

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2159))

Abstract

A common feature of dynamin-related proteins (DRPs) is their use of guanosine triphosphate (GTP) to control protein dynamics. In the case of the endoplasmic- reticulum- (ER)-resident membrane protein atlastin (ATL), GTP binding and hydrolysis result in membrane fusion of ER tubules and the generation of a branched ER network. In this chapter, we describe two independent methods for dissecting the mechanism underlying nucleotide-dependent quaternary structure and conformational changes of ATL, focusing on size-exclusion chromatography coupled with multi-angle light scattering (SEC–MALS) and Förster resonance energy transfer (FRET), respectively. The high temporal resolution of the FRET-based assays enables the ordering of the molecular events identified in structural and equilibrium-based SEC–MALS studies. In combination, these complementary methods report on the oligomeric states of a system at equilibrium and timing of key steps along the enzyme’s catalytic cycle. These methods are broadly applicable to proteins that undergo ligand-induced dimerization and/or conformational changes.

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Acknowledgement

This work was supported by the Spastic Paraplegia Foundation to H.S.

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Author notes

  1. John P. O’Donnell

    Present address: Cell Biology Division, MRC Laboratory of Molecular Biology, Cambridge, UK

  2. John P. O’Donnell and Carolyn M. Kelly contributed equally to this work.

Authors and Affiliations

  1. Department of Molecular Medicine, College of Veterinary Medicine, Cornell University, Ithaca, NY, USA

    John P. O’Donnell, Carolyn M. Kelly & Holger Sondermann

Authors
  1. John P. O’Donnell
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  2. Carolyn M. Kelly
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  3. Holger Sondermann
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Corresponding author

Correspondence to Holger Sondermann .

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Editors and Affiliations

  1. School of Medicine, Case Western Reserve University, Cleveland, OH, USA

    Rajesh Ramachandran

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O’Donnell, J.P., Kelly, C.M., Sondermann, H. (2020). Nucleotide-Dependent Dimerization and Conformational Switching of Atlastin. In: Ramachandran, R. (eds) Dynamin Superfamily GTPases. Methods in Molecular Biology, vol 2159. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0676-6_8

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  • DOI: https://doi.org/10.1007/978-1-0716-0676-6_8

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-0675-9

  • Online ISBN: 978-1-0716-0676-6

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