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Aix Marseille Univ, Centrale Marseille

2 ARTICLES PUBLISHED IN JoVE

Biochemistry

Purification and Quality Control of Recombinant Septin Complexes for Cell-Free Reconstitution

Gerard Castro-Linares¹, Jeffrey den Haan¹, Francois Iv², Carla Silva Martins², Aurélie Bertin³, Manos Mavrakis², Gijsje H. Koenderink¹

¹Department of Bionanoscience, Kavli Institute of Nanoscience Delft, Delft University of Technology, ²Institut Fresnel, CNRS UMR7249, Aix Marseille Univ, Centrale Marseille, ³Institut Curie, Université PSL, Sorbonne Université

In vitro reconstitution of cytoskeletal proteins is a vital tool to understand the basic functional properties of these proteins. The present paper describes a protocol to purify and assess the quality of recombinant septin complexes, which play a central role in cell division and migration.

Biology

Bottom-Up In Vitro Methods to Assay the Ultrastructural Organization, Membrane Reshaping, and Curvature Sensitivity Behavior of Septins

Brieuc Chauvin^*1, Koyomi Nakazawa^*1, Alexandre Beber^1,7, Aurélie Di Cicco¹, Bassam Hajj¹, François Iv², Manos Mavrakis², Gijsje H. Koenderink³, João T. Cabral⁴, Michaël Trichet⁵, Stéphanie Mangenot^*6, Aurélie Bertin^*1

¹Laboratoire Physico Chimie Curie, Institut Curie, PSL Research University, Sorbonne Université, ²Institut Fresnel, CNRS UMR7249, Aix Marseille Univ, Centrale Marseille, ³Department of Bionanoscience, Kavli Institute of Nanoscience Delft, Delft University of Technology, ⁴Department of Chemical Engineering, Imperial College London, ⁵Sorbonne Université, CNRS, Institut de Biologie Paris-Seine (IBPS), Service de microscopie électronique (IBPS-SME), ⁶Laboratoire Matière et Systèmes Complexes (MSC), Université Paris Cité, ⁷Institute of Biotechnology, Czech Academy of Sciences, BIOCEV

Septins are cytoskeletal proteins. They interact with lipid membranes and can sense but also generate membrane curvature at the micron scale. We describe in this protocol bottom-up in vitro methodologies for analyzing membrane deformations, curvature-sensitive septin binding, and septin filament ultrastructure.