Biomedical subjects

Hervé-W Rémigy

Publications and source records attributed to Hervé-W Rémigy.

5 recordsLinked to original sources

PubMed2006

Controlled 2D crystallization of membrane proteins using methyl-beta-cyclodextrin.

High-resolution structural data of membrane proteins can be obtained by studying 2D crystals by electron crystallography. Finding the right conditions to produce these crystals is one of the major bottlenecks encountered in 2D crystallography. Many reviews address 2D crystallization techniques in attempts to provide guidelines for crystallographers. Several techniques including new approaches to remove detergent like the biobeads technique and the development of dedicated devices have been described (dialysis and dilution machines). In addition, 2D crystallization at interfaces has been studied, the most prominent method being the 2D crystallization at the lipid monolayer. A new approach based on detergent complexation by cyclodextrins is presented in this paper. To prove the ability of cyclodextrins to remove detergent from ternary mixtures (lipid, detergent and protein) in order to get 2D crystals, this method has been tested with OmpF, a typical beta-barrel protein, and with SoPIP2;1, a typical alpha-helical protein. Experiments over different time ranges were performed to analyze the kinetic effects of detergent removal with cyclodextrins on the formation of 2D crystals. The quality of the produced crystals was assessed with negative stain electron microscopy, cryo-electron microscopy and diffraction. Both proteins yielded crystals comparable in quality to previous crystallization reports.

Aquaporins↗

PubMed2005

A novel method for detergent concentration determination.

A fast and precise method for detergent concentration determination is presented. (Patent applications for the method described here have been submitted (EP05011904 and US60/702,261). Depending on the interest of the scientific community, the system will be commercialized. (For further information contact Hervé-W. Rémigy at the e-mail address below.) A small droplet of the detergent solution is deposited on a piece of Parafilm M and side views are recorded by two orthogonally arranged TV cameras. The droplet contours are then approximated by ellipses to determine the contact angles. Comparison of the observed contact angle values to calibrated standard curves of known detergent concentrations gives the concentration of the detergent assessed. A range of commonly used detergents was studied to demonstrate the reproducibility and precision of this simple method. As a first application, the detergent binding capacity of the Escherichia coli galactose/proton symporter (GalP) was assessed. Aggregation of GalP was observed when <260 +/- 5 dodecyl-beta,D-maltoside molecules were bound to one GalP molecule. These measurements document the efficacy of the drop-shape based detergent concentration determination described.

Biophysics↗

PubMed2004

Isolation and Characterization of an Outer Membrane Protein of Chlorobium tepidum.

A protein was isolated from membranes of the green sulfur bacterium Chlorobium tepidum. This protein was characterized by gel electrophoresis, gel filtration, analytical ultracentrifugation and amino acid sequencing. The molecular weight of the purified protein was shown to be 26 kDa by SDS-PAGE. HPLC gelfiltration, SDS-PAGE and analytical ultracentrifugation are consistent with the presence of a homogenous protein in the preparations. Amino acid analysis was obtained from the isolated protein after fragmentation with Lys-C, trypsin and cyanogen bromide. The cleavage pattern resulting from these treatments combined with Edman sequencing yield a sequence allowing the identification of an integral membrane agglutinin in Chl. tepidum.

Chlorobium tepidum↗

PubMed2003

Characterization by Mass Spectroscopy of a 10 kDa c-554 Cytochrome from the Green Sulfur Bacterium Chlorobium tepidum.

Preparative isoelectric focusing was used to isolate a type c cytochrome from photosynthetic membranes of the green sulfur bacterium Chlorobium tepidum. The purified protein showed a molecular weight of 10 kDa according to SDS-PAGE and ESI mass spectrometry. The absorption spectrum in the visible range is typical of a cytochrome with peaks at 420, 525.2 and 554.4 nm. Cleavage by either trypsin or endoproteinase lys-C of the isolated cytochrome combined with tandem mass spectrometry and Edman sequencing yielded a sequence perfectly matching parts of the recently sequenced genome of C. tepidum.

Journal Article↗

PubMed2002

The reaction centre from green sulphur bacteria: progress towards structural elucidation.

The reaction centre (RC) of green sulphur bacteria is a FeS-type RC, as are the RCs of Photosystems I (PS I) of oxygenic photosynthetic organisms and of heliobacteria. The core domains of both green sulphur bacterial and heliobacterial RCs are considered to be homodimeric, in contrast to those of purple bacteria, PS I and Photosystem II (PS II). This paper briefly describes the techniques of electron microscopy and image processing suited to investigate the structure of these proteins. Recent advances in the study of the structure of the green sulphur bacterial RC, primarily achieved by the application of scanning transmission electron microscopy, are reviewed.

Journal Article↗