Advanced

Perdeuteration of cholesterol for neutron scattering applications using recombinant Pichia pastoris

Moulin, Martine ; Strohmeier, Gernot A ; Hirz, Melanie ; Thompson, Katherine C ; Rennie, Adrian R ; Campbell, Richard A LU ; Pichler, Harald ; Maric, Selma LU ; Forsyth, V Trevor and Haertlein, Michael (2018) In Chemistry and Physics of Lipids 212. p.80-87
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... (More)

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.

(Less)
Please use this url to cite or link to this publication:
author
; ; ; ; ; ; ; ; and
publishing date
type
Contribution to journal
publication status
published
in
Chemistry and Physics of Lipids
volume
212
pages
8 pages
publisher
Elsevier
external identifiers
  • scopus:85040311890
  • pmid:29357283
ISSN
0009-3084
DOI
10.1016/j.chemphyslip.2018.01.006
language
English
LU publication?
no
id
b81b8c3f-2327-4219-bf2a-83af6842c2f5
date added to LUP
2018-07-19 12:00:46
date last changed
2021-03-31 04:48:45
@article{b81b8c3f-2327-4219-bf2a-83af6842c2f5,
  abstract     = {<p>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.</p>},
  author       = {Moulin, Martine and Strohmeier, Gernot A and Hirz, Melanie and Thompson, Katherine C and Rennie, Adrian R and Campbell, Richard A and Pichler, Harald and Maric, Selma and Forsyth, V Trevor and Haertlein, Michael},
  issn         = {0009-3084},
  language     = {eng},
  pages        = {80--87},
  publisher    = {Elsevier},
  series       = {Chemistry and Physics of Lipids},
  title        = {Perdeuteration of cholesterol for neutron scattering applications using recombinant Pichia pastoris},
  url          = {http://dx.doi.org/10.1016/j.chemphyslip.2018.01.006},
  doi          = {10.1016/j.chemphyslip.2018.01.006},
  volume       = {212},
  year         = {2018},
}