Nicolas Garnier

euHCVdb: the European hepatitis C virus database

The hepatitis C virus (HCV) genome shows remarkable sequence variability, leading to the classification of at least six major genotypes, numerous subtypes and a myriad of quasispecies within a given host. A database allowing researchers to investigate the genetic and structural variability of all available HCV sequences is an essential tool for studies on the molecular virology and pathogenesis of hepatitis C as well as drug design and vaccine development. We describe here the European Hepatitis C Virus Database (euHCVdb, ), a collection of computer-annotated sequences based on reference genomes. The annotations include genome mapping of sequences, use of recommended nomenclature, subtyping as well as three-dimensional (3D) molecular models of proteins. A WWW interface has been developed to facilitate database searches and the export of data for sequence and structure analyses. As part of an international collaborative effort with the US and Japanese databases, the European HCV Database (euHCVdb) is mainly dedicated to HCV protein sequences, 3D structures and functional analyses.

Structural characterization of lipid constituents from natural substances preserved in archaeological environments

The development of a research field at the border between analytical chemistry and archaeology, namely biomolecular archaeology, provides new methods for the study of organic remains highly sensitive to natural decay. Using infrared spectroscopy, gas chromatography and mass spectrometry, it is now possible to chemically identify a series of natural substances preserved in archaeological environments. This paper details the amorphous organic residues discovered in ancient pottery or adhering to flint tools and presents an overview of the analytical methodology developed in our laboratories for the characterization of such remains. Various natural products could be identified, such as animal or plant fats, beeswax or birch bark tar, this latter substance being an adhesive made by a controlled heating of white birch bark.

Characterization of thermally assisted hydrolysis and methylation products of polyphenols from modern and archaeological vine derivatives using gas chromatography–mass spectrometry

Abstract If some ceramics, such as vinary amphorae attest the consumption and trade of wine in the Roman world, the first wine productions in Occident often stay undocumented. Chemical analysis of organic materials preserved in archaeological vessels is the only way to bring new lights about the elaboration and the consumption of this fermented beverage. To determine the preservation state of wine and other grape derivatives residues, we proceeded to in situ tetramethylammonium hydroxide (TMAH) treatment followed by thermally assisted hydrolysis and methylation—gas chromatography—mass spectrometry (THM–GC–MS). The objectives of the study were (i) the understanding of the pyrolytic mechanisms of proanthocyanidins and (ii) the assessment of the usefulness of thermochemolysis for the identification of phenolic markers from polymeric solid deposit in modern wine bottle and Roman amphorae. THM–GC–MS was revealed to be an efficient method for the characterization of fruit derivatives even if mixed with another organic material, such as pitch used to ensure the watertightness of the ceramic container. The preservation of tannins during millennia in archaeological context is here enlightened for the first time by using analytical pyrolysis. The proposed identification of vinary residues is now based on the detection of the association of more than 30 pyrolytic markers derived from di- and trimethoxylated benzenoid compounds. THM–GC–MS represents a new method for the rapid detection of wine traces in ancient ceramics, adapted to tiny samples (

Identification of Animal Glue Species in Artworks Using Proteomics: Application to a 18th Century Gilt Sample

This study proposes a proteomic-based strategy for the identification of the origin species of glues used as binding media and adhesives in artworks. The methodology, based on FTICR high resolution mass spectrometry, was evaluated on glues from different animal origin (i.e., bovine, rabbit, and fish). The analysis of the peptide mixture resulting from the enzymatic hydrolysis of the proteins led to the identification of species-specific peptides. Up to 15 specific peptides were identified for the bovine species and three for the rabbit species and, in the case of sturgeon glue, three fish-specific peptides were found by sequence homology to the rainbow trout. Then, the method was applied to authenticate different rabbit skin glue samples, including a 100 year-old sample named Colle à Doreurs coming from the Maison Totin-Frères. For this sample, two specific peptides of rabbit collagen were identified. To evaluate the method in a complex matrix, model paints composed of lead white, linseed oil, and animal glue were prepared. Species-specific peptides were identified in each paint sample. Finally, a gilt sample from St Maximin church dating from the eighteenth century was analyzed, and 13 peptides specific to bovine collagens were identified starting from very low sample amount (50 μg).

Dealing with the identification of protein species in ancient amphorae

This manuscript deals with the identification of protein residues in amphorae, including particularly identification of protein species. The work described was performed on fishes, the anchovy (Engraulis encrasicolus) and bonito (Sarda sarda) species frequently found in the Mediterranean area. Based on proteomic techniques, the analytical strategy was adapted to analysis of protein residues from tiny ceramic fragments. The major difficulty was to extract proteins and limit their hydrolysis during the sample preparation; consequently, multiple soft extraction techniques were evaluated. The most valuable results were obtained using a solution containing high amounts of denaturing agents, urea and thiourea, reducing agent, dithiothreitol, and detergent, 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate. The analysis using nano liquid chromatography–nano electrospray ionization double quadrupole time-of-flight mass spectrometry resulted in the identification of up to 200 proteins for the anchovy and bonito species, among which 73 peptides were found to be fish-specific. Because bonito and anchovy species are not documented and fully sequenced in genomic databases, the preliminary protein identification was realized via sequence homology to other fish sequenced species. Amino acid substitutions of peptides were assigned on the basis of the interpretation of tandem mass spectrometry spectra using de novo sequencing; these peptides, not reported up to now in databases, constitute species-specific markers. The method developed was finally applied to an archaeological sample replica impregnated with a mixture of fish tissue from both species; this experiment successfully led to the identification of 17 fish proteins, including 33 fish-specific peptides. This work shows that the analytical method developed has great potential for the identification of protein species in complex archaeological samples.

Nonlinear dynamics of waves and modulated waves in 1D thermocapillary flows. I. General presentation and periodic solutions

Abstract We present experimental results on hydrothermal traveling waves dynamics in long and narrow 1D channels. The onset of primary traveling-wave patterns is briefly presented for different fluid heights and for annular or bounded channels, i.e., within periodic or non-periodic boundary conditions. For periodic boundary conditions, by increasing the control parameter or changing the discrete mean wavenumber of the waves, we produce modulated wave patterns. These patterns range from stable periodic phase-solutions, due to supercritical Eckhaus instability, to spatio-temporal defect-chaos involving traveling holes and/or counter-propagating waves competition, i.e., traveling sources and sinks. The transition from non-linearly saturated Eckhaus modulations to transient pattern breaks by traveling holes and spatio-temporal defects is documented. Our observations are presented in the framework of coupled complex Ginzburg–Landau equations with additional fourth and fifth order terms which account for the reflection symmetry breaking at high wave-amplitude far from onset. The second part of this paper [N. Garnier, A. Chiffaudel, F. Daviaud, Nonlinear dynamics of waves and modulated waves in 1D thermocapillary flows. II. Convective/absolute transitions, Physica D (2003), this issue] extends this study to spatially non-periodic patterns observed in both annular and bounded channel.

Automated method for analysis of tryptophan and tyrosine metabolites using capillary electrophoresis with native fluorescence detection.

Capillary electrophoresis (CE) with laser-induced native fluorescence (LINF) detection offers the ability to characterize low levels of selected analyte classes, depending on the excitation and emission wavelengths used. Here a new automated CE-LINF system that provides deep ultraviolet (DUV) excitation (224xa0nm) and variable emission wavelength detection was evaluated for the analysis of small molecule tryptophan- and tyrosine-related metabolites. The optimized instrument design includes several features that increase throughput, lower instrument cost and maintenance, and decrease complexity when compared with earlier systems using DUV excitation. Sensitivity is enhanced by using an ellipsoid detection cell to increase the fluorescence collection efficiency. The limits of detection ranged from 4 to 30xa0nmol/L for serotonin and tyrosine, respectively. The system demonstrated excellent linearity over several orders of magnitude of concentration and intraday precision from 1–11xa0% relative standard deviation (RSD). The instrument’s performance was validated via tryptophan and serotonin characterization using tissue extracts from the mammalian brain stem, with RSDs of less than 10xa0% for both metabolites. The flexibility and sensitivity offered by DUV laser excitation and tunable emission enables a broad range of small-volume measurements.

Nonlinear dynamics of waves and modulated waves in 1D thermocapillary flows. II. Convective/absolute transitions

Abstract We present experimental results on hydrothermal waves in long and narrow 1D channels. In a bounded channel, we describe the primary and secondary instabilities leading to waves and modulated waves in terms of convective/absolute transitions. Because of on the combined effect of finite group velocity and of the presence of boundaries, the wave patterns are non-uniform in space. We also investigate non-uniform wave patterns observed in an annular channel in the presence of sources and sinks of hydrothermal waves. We connect our observations with the complex Ginzburg–Landau model equation in the very same way as in the first part of the paper [Nonlinear dynamics of waves and modulated waves in 1D thermocapillary flows. I. General presentation and periodic solutions, Physica D (2003), this issue].

SM2PH‐db: an interactive system for the integrated analysis of phenotypic consequences of missense mutations in proteins involved in human genetic diseases

Understanding how genetic alterations affect gene products at the molecular level represents a first step in the elucidation of the complex relationships between genotypic and phenotypic variations, and is thus a major challenge in the postgenomic era. Here, we present SM2PH‐db (http://decrypthon.igbmc.fr/sm2ph), a new database designed to investigate structural and functional impacts of missense mutations and their phenotypic effects in the context of human genetic diseases. A wealth of up‐to‐date interconnected information is provided for each of the 2,249 disease‐related entry proteins (August 2009), including data retrieved from biological databases and data generated from a Sequence–Structure–Evolution Inference in Systems‐based approach, such as multiple alignments, three‐dimensional structural models, and multidimensional (physicochemical, functional, structural, and evolutionary) characterizations of mutations. SM2PH‐db provides a robust infrastructure associated with interactive analysis tools supporting in‐depth study and interpretation of the molecular consequences of mutations, with the more long‐term goal of elucidating the chain of events leading from a molecular defect to its pathology. The entire content of SM2PH‐db is regularly and automatically updated thanks to a computational grid data federation facilities provided in the context of the Decrypthon program. Hum Mutat 31:127–135, 2010.

Blast sampling for structural and functional analyses

BackgroundThe post-genomic era is characterised by a torrent of biological information flooding the public databases. As a direct consequence, similarity searches starting with a single query sequence frequently lead to the identification of hundreds, or even thousands of potential homologues. The huge volume of data renders the subsequent structural, functional and evolutionary analyses very difficult. It is therefore essential to develop new strategies for efficient sampling of this large sequence space, in order to reduce the number of sequences to be processed. At the same time, it is important to retain the most pertinent sequences for structural and functional studies.ResultsAn exhaustive analysis on a large scale test set (284 protein families) was performed to compare the efficiency of four different sampling methods aimed at selecting the most pertinent sequences. These four methods sample the proteins detected by BlastP searches and can be divided into two categories: two customisable methods where the user defines either the maximal number or the percentage of sequences to be selected; two automatic methods in which the number of sequences selected is determined by the program. We focused our analysis on the potential information content of the sampled sets of sequences using multiple alignment of complete sequences as the main validation tool. The study considered two criteria: the total number of sequences in BlastP and their associated E-values. The subsequent analyses investigated the influence of the sampling methods on the E-value distributions, the sequence coverage, the final multiple alignment quality and the active site characterisation at various residue conservation thresholds as a function of these criteria.ConclusionThe comparative analysis of the four sampling methods allows us to propose a suitable sampling strategy that significantly reduces the number of homologous sequences required for alignment, while at the same time maintaining the relevant information concerning the active site residues.