Year: 2020
Type: article
Abstract: Today, the sedimentation of proteins into a magic-angle spinning (MAS) rotor gives access to fast and reliable sample preparation for solid-state Nuclear Magnetic Resonance (NMR), and this has allowed for the investigation of a variety of non-crystalline protein samples. High protein concentrations on the order of 400 mg/mL can be achieved, meaning that around 50-60% of the NMR rotor content is protein; the rest is a buffer solution, which includes counter ions to compensate for the charge of the protein. We have demonstrated herein the long-term stability of four sedimented proteins and complexes thereof with nucleotides, comprising a bacterial DnaB helicase, an ABC transporter, an archaeal primase, and an RNA polymerase subunit. Solid-state NMR spectra recorded directly after sample filling and up to 5 years later indicated no spectral differences and no loss in signal intensity, allowing us to conclude that protein sediments in the rotor can be stable over many years. We have illustrated, using an example of an ABC transporter, that not only the structure is maintained, but that the protein is still functional after long-term storage in the sedimented state.
Language: English
Subfield: Spectroscopy
Field: Chemistry
Domain: Physical Sciences
Open Access status: gold
Estimated APC paid: $2,001
Funders: National Science Foundation, European Commission, Agence Nationale de la Recherche, Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung, Eidgenössische Technische Hochschule Zürich, Université de Lyon, Laboratoire d'Excellence Ecofect, European Research Council, H2020 European Research Council
Awards: G169068394, G2125209779, Structure and Function of a Primase (prolongation), PROJET AVENIR LYON SAINT-ETIENNE, G3010527862, G36366808, G3785575267, G434337789, Zooming in on the conformational details of a membrane efflux pump, NMR studies in the Solid State, Dynamiques eco-évolutives des maladies infectieuses, G4866930200, Faster magic-angle spinning leads to a resolution revolution in biological solid-state NMR, G5649854608, G628552264, G7835500200, Dissecting the molecular mechanism of an antibiotic efflux pump, G8690818240, G8886589746, G9575639
OpenAlex ID: W3007801938