Moulin, M;
Strohmeier, GA;
Hirz, M;
Thompson, KC;
Rennie, AR;
Campbell, RA;
Pichler, H;
... Haertlein, M; + view all
(2018)
Perdeuteration of cholesterol for neutron scattering applications using recombinant Pichia pastoris.
Chemistry and Physics of Lipids
, 212
pp. 80-87.
10.1016/j.chemphyslip.2018.01.006.
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Abstract
Deuteration of biomolecules has a major impact on both quality and scope of neutron scattering experiments. Cholesterol is a major component of mammalian cells, where it plays a critical role in membrane permeability, rigidity and dynamics, and contributes to specific membrane structures such as lipid rafts. Cholesterol is the main cargo in low and high-density lipoprotein complexes (i.e. LDL, HDL) and is directly implicated in several pathogenic conditions such as coronary artery disease which leads to 17 million deaths annually. Neutron scattering studies on membranes or lipid-protein complexes exploiting contrast variation have been limited by the lack of availability of fully deuterated biomolecules and especially perdeuterated cholesterol. The availability of perdeuterated cholesterol provides a unique way of probing the structural and dynamical properties of the lipoprotein complexes that underly many of these disease conditions. Here we describe a procedure for in vivo production of perdeuterated recombinant cholesterol in lipid-engineered Pichia pastoris using flask and fed-batch fermenter cultures in deuterated minimal medium. Perdeuteration of the purified cholesterol was verified by mass spectrometry and its use in a neutron scattering study was demonstrated by neutron reflectometry measurements using the FIGARO instrument at the ILL.
| Type: | Article |
|---|---|
| Title: | Perdeuteration of cholesterol for neutron scattering applications using recombinant Pichia pastoris |
| Open access status: | An open access version is available from UCL Discovery |
| DOI: | 10.1016/j.chemphyslip.2018.01.006 |
| Publisher version: | https://doi.org/10.1016/j.chemphyslip.2018.01.006 |
| Language: | English |
| Additional information: | Copyright © 2018 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY license (http://creativecommons.org/licenses/BY/4.0/). |
| Keywords: | Perdeuteration, Cholesterol, Pichia pastoris, Neutron scattering, Lipid engineering |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Div of Biosciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Div of Biosciences > Structural and Molecular Biology |
| URI: | https://discovery.ucl.ac.uk/id/eprint/10073381 |
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