Characterizing a monotopic membrane enzyme. Biochemical, enzymatic and crystallization studies on Aquifex aeolicus sulfide:quinone oxidoreductase

Monotopic membrane proteins are membrane proteins that interact with only one leaflet of the lipid bilayer and do not possess transmembrane spanning segments. They are endowed with important physiological functions but until now only few of them have been studied. Here we present a detailed biochemical, enzymatic and crystallographic characterization of the monotopic membrane protein sulfide:quinone oxidoreductase. Sulfide:quinone oxidoreductase is a ubiquitous enzyme involved in sulfide detoxification, in sulfide-dependent respiration and photosynthesis, and in heavy metal tolerance. It may also play a crucial role in mammals, including humans, because sulfide acts as a neurotransmitter in these organisms. We isolated and purified sulfide:quinone oxidoreductase from the native membranes of the hyperthermophilic bacterium Aquifex aeolicus. We studied the pure and solubilized enzyme by denaturing and non-denaturing polyacrylamide electrophoresis, size-exclusion chromatography, cross-linking, analytical ultracentrifugation, visible and ultraviolet spectroscopy, mass spectrometry and electron microscopy. Additionally, we report the characterization of its enzymatic activity before and after crystallization. Finally, we discuss the crystallization of sulfide:quinone oxidoreductase in respect to its membrane topology and we propose a classification of monotopic membrane protein crystal lattices. Our data support and complement an earlier description of the three-dimensional structure of A. aeolicus sulfide:quinone oxidoreductase (M. Marcia, U. Ermler, G. Peng, H. Michel, Proc Natl Acad Sci USA, 106 (2009) 9625-9630) and may serve as a reference for further studies on monotopic membrane proteins. (C) 2010 Elsevier B.V. All rights reserved.

Investigation into the rheological properties of PES/NMP/nonsolvent membrane-forming systems

From the perspective of a polymer solution, the rheological properties of the popular polyethersulfone (PES)/N-methyl-2-pyrrolidone (NMP)/nonsolvent (NS) membrane-forming system were investigated thoroughly with a controlled stress rheometer (HAAKE RS75, Germany). The scope of the study included measurements of the controlled-stress flow curve, creep recovery, and dynamic oscillation. H2O, 1-butanol, ethylene glycol, and diethylene glycol were used as NS additives. The effects of the polymer concentration and the quality of the solvent mixture, as characterized by the approaching ratio, on the rheological behavior of the dopes were studied. Up to 38 wt % PES and extremely adjacent to the phase separation (i.e., the approaching ratio of the dope was 0.95), the viscous property dominated all the dopes, which behaved as Newtonian fluids. Moreover, all the membrane-forming dopes investigated were in the crossover regime in the semidilute region, in which the chains were overlapping but unentangled. (C) 2001 John Wiley & Sons, Inc.

Modification of nylon membrane used for affinity adsorption

Nylon membrane was modified by binding with polyhydroxyl-containing materials to increase its hydrophilicity and reduce its nonspecific interaction with proteins. The effect of binding hydrophilic materials on amount of ligand bound-Cibacron Blue F3GA (CBF) was investigated. Experimental data showed that the amount of CBF bound can be increased significantly after binding of hydrophilic materials.

Removal of endotoxin from human serum albumin solutions by hydrophobic and cationic charged membrane

A novel matrix of macropore cellulose membrane was prepared by chemical graft, and immobilized the cationic charged groups as affinity ligands. The prepared membrane Fan be used for the removal of endotoxin from human serum albumin (HSA) solutions. With a cartridge of 20 sheets affinity membrane of 47 mm diameter, the endotoxin level in HSA solution can be reduced ro 0.027 eu/mL. Recovery of HSA was over 95%.

Partial oxidation of ethane to syngas in an oxygen-permeable membrane reactor

A perovskite-type oxide of Ba0.5Sr0.5Co0.8Fe0.2O3-delta (BSCFO) with mixed electronic and oxygen ionic conductivity at high temperatures was used as an oxygen-permeable membrane. A tubular membrane of BSCFO made by extrusion method has been used in the membrane reactor to exclusively transport oxygen for the partial oxidation of ethane (POE) to syngas with catalyst of LiLaNiO/gamma-Al2O3 at temperatures of 800-900 degreesC. After only 30 min POE reaction in the membrane reactor, the oxygen permeation flux reached at 8.2 ml cm(-2) min(-1). After that, the oxygen permeation flux increased slowly and it took 12 h to reach at 11.0 ml cm(-2) min(-1). SEM and EDS analysis showed that Sr and Ba segregations occurred on the used membrane surface exposed to air while Co slightly enriched on the membrane surface exposed to ethane. The oxygen permeation flux increased with increasing of concentration of C2H6, which was attributed to increasing of the driving force resulting from the more reducing conditions produced with an increase of concentration of C2H6 in the feed gas. The tubular membrane reactor was successfully operated for POE reaction at 875 degreesC for more than 100 h without failure, with ethane conversion of similar to 100%, CO selectivity of >91% and oxygen permeation fluxes of 10-11 ml cm(-2) min(-1). (C) 2002 Elsevier Science B.V. All rights reserved.

Alkaline treatment of the cellulose fiber affecting membrane column behaviour for high-performance immunoaffinity chromatography

The original cellulose fibers and those treated by alkaline solution were both used to prepare the acrylic membranes. The two kinds of membranes were packed into the columns for high-performance immunoaffinity chromatography by the immobilization of protein A on them. It was observed that the alkaline treatment of the cellulose fiber decreased the pressure resistance of the membrane to the mobile phases and greatly increased the accessible volume to the proteins, but affected the adsorption capacity of human IgG on the protein A membrane columns less. There is little difference between those two kinds of membranes on the adsorption capacities of HIgG, which means that the alkaline treatment of the cellulose fiber only significantly changes the void volume inter-membrane, and the porosity and surface area of membrane less. Alkaline treatment of the cellulose fiber reduced the membrane-column efficiency significantly. Some typical examples for the immunoaffinity analysis of IgG from human and dog plasma on the protein A membrane columns are illustrated. Copyright (C) 2000 John Wiley & Sons, Ltd.

Syngas production using an oxygen-permeating membrane reactor with cofeed of methane and carbon dioxide

CH4-CO2-O-2 reforming to syngas in a never Ba0.5Sr0.5Co0.8Fe0.2O3.delta oxygen-permeable membrane reactor using LiLaNiO/gamma-Al2O3 as catalyst was successfully reported. Excellent reaction performance was achieved with around 92% methane conversion efficiency, 95% CO2 conversion rate, and nearly 8.5mL/min.cm(2) oxygen permeation flux. In contrast to the oxygen permeation model with the presence of large concentration of CO2 (under such condition the oxygen permeation flux deteriorates with time), the oxygen permeation flux is really stable under the CH4CO2-O-2 reforming condition.