Perdeuteration of cholesterol for neutron scattering applications using recombinant Pichia pastoris

Deuteration of biomolecules has a great 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 using neutron reflectometry

Deuterated detergents for structural and functional studies of membrane proteins: Properties, chemical synthesis and applications

Detergents are amphiphilic compounds that have crucial roles in the extraction, purification and stabilization of integral membrane proteins and in experimental studies of their structure and function. One technique that is highly dependent on detergents for solubilization of membrane proteins is solution-state NMR spectroscopy, where detergent micelles often serve as the best membrane mimetic for achieving particle sizes that tumble fast enough to produce high-resolution and high-sensitivity spectra, although not necessarily the best mimetic for a biomembrane. For achieving the best quality NMR spectra, detergents with partial or complete deuteration can be used, which eliminate interfering proton signals coming from the detergent itself and also eliminate potential proton relaxation pathways and strong dipole-dipole interactions that contribute line broadening effects. Deuterated detergents have also been used to solubilize membrane proteins for other experimental techniques including small angle neutron scattering and single-crystal neutron diffraction and for studying membrane proteins immobilized on gold electrodes. This is a review of the properties, chemical synthesis and applications of detergents that are currently commercially available and/or that have been synthesized with partial or complete deuteration. Specifically, the detergents are sodium dodecyl sulphate (SDS), lauryldimethylamine-oxide (LDAO), n-octyl-?-D-glucoside (?-OG), n-dodecyl-?-D-maltoside (DDM) and fos-cholines including dodecylphosphocholine (DPC). The review also considers effects of deuteration, detergent screening and guidelines for detergent selection. Although deuterated detergents are relatively expensive and not always commercially available due to challenges associated with their chemical synthesis, they will continue to play important roles in structural and functional studies of membrane proteins, especially using solution-state NMR

Wholesale pricing in a small open economy

This paper addresses the empirical analysis of wholesale profit margins using data of the Dutch wholesale sector, 1986. At the heart of the analysis is the typical nature of wholesale production: wholesalers do not produce a tangible product, but offer a service capacity. This has an immediate impact on the identification, interprelation and measurement of determinants of profit variations. A model is set up to explain variations in wholesale profit margins, which is inspired by two widely applied approaches to industry pricing: the behavioural mark-up model and the marginalist price-cost model

Inter-industry Variations in the Effect of Trade on Industry Performance.

This paper develops an empirical model to measure the strength of import discipline across industries, allowing it to vary sys tematically with various features of industry market structure. The m odel is applied to a panel of data for eighty-two Belgian three-digit industries for the period 1973-78, and the results suggest that the parameters of the structural model are stable over time, that the ext ent of discipline is endogenous, and that about 70 percent of the ind ustries in the sample experience significant trade-induced competitiv e discipline. Copyright 1988 by Blackwell Publishing Ltd.

THE FANTASIO SET-UP (I): DESCRIPTION AND EXTENSION TOWARDS FEMTO-FANTASIO

M. Herman, K. Didriche, D. Hurtmans, B. Kizil, P. Macko, A. Rizopoulos and P. Van Poucke, Mol. Phys., \underline{105Author Institution: Service de Chimie quantique et Photophysique CP160/09, Faculte des Sciences, Universite Libre de; Bruxelles (U.L.B.), Av. Roosevelt, 50, B-1050, Bruxelles, Belgium; Laboratoire de; Spectrometrie Physique, Laboratoire de Spectrométrie Physique, Universite Joseph Fourier de; Grenoble, B.P. 87, 38402 Saint-Martin-d'Heres Cedex, FranceWe have built the FANTASIO set-up (for "Fourier trANsform, Tunable diode and quadrupole mAss spectrometers interfaced to a Supersonic expansIOn") to investigate jet-cooled molecules and dimers, 843 (2007). }. The set-up will be described. New developments are on the way, to be illustrated by preliminary results. These include the doubling of the vacuum pumping system efficiency. They will also allow for a Femto OPO system to be used as a broadband, cavity enhanced absorption source to be interfaced to the supersonic expansion and to the high resolution FTIR instrument of FANTASIO