Medium chain length polyhydroxyalkanoates (mcl-PHA) have been produced by fed-batch cultivation o... more Medium chain length polyhydroxyalkanoates (mcl-PHA) have been produced by fed-batch cultivation of Pseudomonas oleovorans. Using octanoic acid as the sole carbon substrate, a copolymer of 3-hydroxyoctanoate and 3-hydroxyhexanoate monomer units was accumulated up to 38 wt. % of cell mass. Mcl-PHA based nanocomposites have then been prepared using Cloisite 15A as nanofiller and characterized. Wide angle X-ray diffraction and rheological analysis of the nanocomposites highlighted their morphologies, which can be considered as well-intercalated structures. The thermal characteristics of mcl-PHA are not significantly affected by the nanofiller incorporation. On the contrary, the storage moduli of nanocomposites are greatly enhanced with increasing clay fraction due to the degree of dispersion of the nanoplatelets in the polymer matrix.
Medium chain length polyhydroxyalkanoates (mcl-PHA) have been produced by fed-batch cultivation o... more Medium chain length polyhydroxyalkanoates (mcl-PHA) have been produced by fed-batch cultivation of Pseudomonas oleovorans. Using octanoic acid as the sole carbon substrate, a copolymer of 3-hydroxyoctanoate and 3-hydroxyhexanoate monomer units was accumulated up to 38 wt. % of cell mass. Mcl-PHA based nanocomposites have then been prepared using Cloisite 15A as nanofiller and characterized. Wide angle X-ray diffraction and rheological analysis of the nanocomposites highlighted their morphologies, which can be considered as well-intercalated structures. The thermal characteristics of mcl-PHA are not significantly affected by the nanofiller incorporation. On the contrary, the storage moduli of nanocomposites are greatly enhanced with increasing clay fraction due to the degree of dispersion of the nanoplatelets in the polymer matrix.
The Journal of Steroid Biochemistry and Molecular Biology, 2015
Estrogens are steroid hormones that play a pivotal role in growth, differentiation and function o... more Estrogens are steroid hormones that play a pivotal role in growth, differentiation and function of reproductive and non-reproductive tissues, mediated through estrogen receptors (ERs). Estrogens are involved in different genomic and non-genomic cell signaling pathways which involve well-defined subcellular ER localizations. Thus, ER activity results from complex interplays between intrinsic binding properties and specific subcellular localization. Since these two factors are deeply intricate, we carried out, in a unique yeast cell context, a comparative study to better understand structure/function/ subcellular distribution relationships. This was carried out by comparing two ERs: the human ER a subtype (hERa) and the short form of the a isoform of the rainbow trout ER (rtERa S ). Their distinct binding properties to agonist and antagonist ligands and subcellular localizations were characterized in Saccharomyces cerevisiae yeast cells. An unexpected partial agonistic effect of ICI 182-780 was observed for rtERa S . Concomitant to distinct binding properties, distinct subcellular localizations were observed before and after ligand stimulation. Due to the unique cell context, the link between ERs intrinsic binding properties and subcellular localizations is partly unveiled and issues are hypothesized based on the role of cytoplasmic transient complexes which play a role in the ER cytoplasmic/nuclear partition, which in turn is critical for the recruitment of co-regulators in the nucleus.
A high-pressure Fourier-transform infrared technique was used to probe the evolution of 'H bonds ... more A high-pressure Fourier-transform infrared technique was used to probe the evolution of 'H bonds inside the helical segments of myoglobin in relation to p2H, Tris concentration in the medium and iron-ligand nature. The analysis was focused on changes in the conformation-sensitive amide-I' band, reflecting the peptide C = 0 group stretching vibrations coupled to the in-plane N-2H bending and C = N stretching modes. From data obtained under high pressure, the strength of 2H bonds, inside the a-helical segments of the protein at atmospheric pressure, is not simply a function of p2H and salt concentration. At low Tris concentration (50 mM), the strength of these 2H bonds increases with p'H, whereas for a higher Tris concentration (100 mM) this strength is lower at p2H 7 than at p'H 6.0 or 8.5. It is also observed that the azidometmyoglobin molecule exhibits tighter intrahelical interactions and lower sensitivity to pressure than aquametmyoglobin. Information is also presented regarding interhelical interactions in relation to the solvent.
Biochimica et Biophysica Acta (BBA) - Lipids and Lipid Metabolism, 1985
GalN induced a rapid inhibition of prelabelled triacylglycerol secretion by hepatocytes isolated ... more GalN induced a rapid inhibition of prelabelled triacylglycerol secretion by hepatocytes isolated from fed rats in which this secretion occurred to a larger extent than in hepatocytes isolated from fasted rats. The data reported here suggest that GalN induces a rise of triacylglycerol synthesis by inhibiting the palmitate oxidation pathway and a decrease of triacylglycerol secretion through an early derangement of the secretory pathway.
Chalcogenide glass optical fibers possess very low optical losses in the middle infrared range fr... more Chalcogenide glass optical fibers possess very low optical losses in the middle infrared range from 2 to 12 mm. They were used to implement remote infrared spectroscopy, known as Fiber Evanescent Wave Spectroscopy (FEWS). Due to their hydrophobic behavior, such sensor is especially suitable for application in biology and medicine where water is a nuisance to detect relevant information. Moreover,
Chalcogenide glasses are a matchless material as far as mid-infrared (IR) applications are concer... more Chalcogenide glasses are a matchless material as far as mid-infrared (IR) applications are concerned. They transmit light typically from 2 to 12 μm and even as far as 20 μm depending on their composition, and numerous glass compositions can be designed for optical fibers. One of the most promising applications of these fibers consists in implementing fiber evanescent wave spectroscopy, which enables detection of the mid-IR signature of most biomolecules. The principles of fiber evanescent wave spectroscopy are recalled together with the benefit of using selenide glass to carry out this spectroscopy. Then, two large-scale studies in recent years in medicine and food safety are exposed. To conclude, the future strategy is presented. It focuses on the development of rare earth-doped fibers used as mid-IR sources on one hand and tellurium-based glasses to shift the limit of detection toward longer wavelength on the other hand.
For decades, the origin of the concentric ring pattern of bacterial swarming colonies has puzzled... more For decades, the origin of the concentric ring pattern of bacterial swarming colonies has puzzled microbiologists. It was hypothesized that a periodic water activity variation originates a phase transition within the extracellular matrix water H bond network, which switches on and off the exopolysaccharide auto-organization. Both rheological and infrared spectroscopy measurements respectively performed at a molecular scale and on a currently migrating colony, have given a physical insight into the mechanisms which underlie the switch between swarming and consolidation phases. Thanks to in situ and real time infrared microspectroscopy, and thanks to the brilliance of the infrared beam at SOLEIL synchrotron, here we demonstrate that Proteus mirabilis swarming is triggered by a periodic variation of water activity at the colony edge. A dynamic behavior emerges from the global properties of the multicellular entity which relies on the ability of the bacterial cells to tune exoproduct synthesis in order to undergo sharp transitions at a given water activity threshold.
The pH dependence of the tertiary structural changes and the quaternary reorganization of the a j... more The pH dependence of the tertiary structural changes and the quaternary reorganization of the a j interface of oxyhemoglobin in solution has been previously observed in our laboratory. The present work aims to establish whether or not these progressive structural changes with pH result from proton exchange between the protein and the solvent. We therefore have used infrared spectroscopy, acidlbase titration and 'HI2H exchange to assess the effect of external proton concentration on the structural and dynamic properties of the hemoglobin molecule in solution. This study is also performed on the carbonmonoxy form since the affinity of hemoglobin for CO is much higher than for oxygen. The present findings demonstrate that affinity changes arc closely related to protein fluctuations, as well as structural modifications. An increased affinity, induced either by ligand replacement or pH variation, is associated with the constrained chains exhibiting a lower degree of fluctuation, together with a looser a/3 interface association It was recently shown that the tertiary and quaternary structures of oxyhemoglobin in solution evolve continuously with pH [l]. This gradual development was probed by observing the SH stretching vibration of two cysteinyl residues: aCysl04 (G11) and /3Cysll2 (G14). These two cysteine residues are known to be strongly H-bonded and very sensitive to slight conformational changes which are undetectable by other techniques [2, 31. The H-bond affects the distance between the sulfur and hydrogen atoms and, consequently, perturbs the SH vibration. The aCysl04 (G11) residue provides detailed information on the tertiary constraints of the G chain a-helices whereas the thiol group of pCysll2 (G14) is sensitive to quaternary structural changes at the tight ap interface Although the functional properties of hemoglobin are known to be pH dependent [5 -81, the changes in interactions inside the protein, induced by the surrounding solvent, are not yet clearly understood. While the affinity of hemoglobin for oxygen increases with pH, the molecular constraints at the x p interface decrease and the intrachain H-bonds of the a chain become stronger [I]. To investigate further the effect of pH on the molecular organization of hemoglobin, the Fouriertransform infrared study of the stretching vibrations of SH was extended to the carbonmonoxy hemoglobin derivative since the partition coefficient between O2 and CO is about 260 for the fully ligated hemoglobin . Furthermore, in order to obtain a more general understanding of the mechanisms correlating the proton concentration in the external solvent with the dynamic properties of these two hemoglobin forms (0, and CO), we have studied their acid/base titration behaviour and their 'H/2H exchange variations with pH. Indeed, many protons of the protein are exchangeable with those of the external solvent [9 -121. Besides, Lumry and Rosenberg
Fiber evanescent wave spectroscopy (FEWS) was used to classify various foodborne pathogens (Liste... more Fiber evanescent wave spectroscopy (FEWS) was used to classify various foodborne pathogens (Listeria monocytogenes, Staphylococcus aureus and Salmonella enteritidis) using chemometric methods such as principal component analysis (PCA) and logistic-partial least squares (logistic-PLS). Spectra were recorded thanks to an IR optical fiber made of chalcogenide glasses. A limited segregation between the healthy and non healthy food has been observed using the PCA method, whereas a clear differentiation between bacteria strains was obtained using the PLS regression. FEWS coupled to chemometric methods is a promising technique for the identification of foodborne pathogens as it could provide a rapid identification and classification of the bacteria.
Chalcogenide glass optical fibers possess very low optical losses in the middle infrared range fr... more Chalcogenide glass optical fibers possess very low optical losses in the middle infrared range from 2 to 12 mm. They were used to implement remote infrared spectroscopy, known as Fiber Evanescent Wave Spectroscopy (FEWS). Due to their hydrophobic behavior, such sensor is especially suitable for application in biology and medicine where water is a nuisance to detect relevant information. Moreover, the design of the sensor using tapered fibers enables to improve the signal to noise ratio. Then, once coupled with unsupervised analysis technique such as Principle Component Analysis (PCA), it has been shown that this tool is efficient to differentiate between obese and control mice by recording their serum FEWS spectra. The same method has been carried out to detect in situ the both phenotypes of a bacterial culture.
Owing to their low losses in the 2-12 micrometers region, the Te- As-Se glass fibers are used for... more Owing to their low losses in the 2-12 micrometers region, the Te- As-Se glass fibers are used for infrared light transportation as well as sensing element based on evanescent wave absorption mechanism. The efficiency of the system is improved by tapering the fiber diameter in the sensing zone. With this kind of sensor, infrared spectra of biological tissue have been obtained. Spectra of mouse liver have been especially recorded in order to detect spectral differences between the healthy and the tumoral states of mouse liver.
Fiber evanescent wave spectroscopy (FEWS) explores the mid-infrared domain, providing information... more Fiber evanescent wave spectroscopy (FEWS) explores the mid-infrared domain, providing information on functional chemical groups represented in the sample. Our goal is to evaluate whether spectral fingerprints obtained by FEWS might orientate clinical diagnosis. Serum samples from normal volunteers and from four groups of patients with metabolic abnormalities are analyzed by FEWS. These groups consist of iron overloaded genetic hemochromatosis (GH), iron depleted GH, cirrhosis, and dysmetabolic hepatosiderosis (DYSH). A partial least squares (PLS) logistic method is used in a training group to create a classification algorithm, thereafter applied to a test group. Patients with cirrhosis or DYSH, two groups exhibiting important metabolic disturbances, are clearly discriminated from control groups with AUROC values of 0.94+/-0.05 and 0.90+/-0.06, and sensibility/specificity of 86/84% and 87/87%, respectively. When pooling all groups, the PLS method contributes to discriminate controls, cirrhotic, and dysmetabolic patients. Our data demonstrate that metabolic profiling using infrared FEWS is a possible way to investigate metabolic alterations in patients.
The effects of pH on the a-subunit tertiary structure and on the alpI interface of the hemoglobin... more The effects of pH on the a-subunit tertiary structure and on the alpI interface of the hemoglobin molecule in aqueous solution are observed using Fourier transform infrared spectroscopy ofSH stretching vibrations from the a104 and 6112 cysteinyl residues.
Previously, we have identified an imperfect estrogen response element (rtERE) in the promoter of ... more Previously, we have identified an imperfect estrogen response element (rtERE) in the promoter of the rainbow trout vitellogenin gene. Although this ERE leads to a lower transcriptional activation, a better estradiol stimulation in vivo as compared to consensus ERE (EREcs) was observed. Here we examine the ability of recombinant human estrogen receptor R (rhERR) to bind DNA containing the EREcs or the natural imperfect rtERE, which contains three mismatches. At low salt concentration, whatever the ERE sequence, dissociation equilibrium constants of the specific rhERR-ERE complexes are similar (K D ) 2 nM) with the same stoichiometry. As salt concentration increases from 80 to 200 mM KCl, the affinity of the rhERR-rtERE complex largely diminishes whereas that of rhERR-EREcs seems less affected. Hence the nature of the interactions stabilizing these complexes is different: more ionic in rhERR-rtERE as compared to rhERR-EREcs. Moreover, kinetic measurements showed that specific rhERR-ERE complexes exhibit shorter half-lives (few seconds) and that the rhERR-EREcs complex is more stable (33 s) than the complex that formed with rtERE (19.8 s), in accordance with equilibrium binding results. Finally, dynamic studies of rhERR have shown that the protein fluctuations are damped when the salt concentration increases or when bound to ERE and all the more with rtERE. The interplay of affinity, complex half-lives, and protein dynamics in the transcriptional regulation of estrogen receptor is discussed.
In this study we investigated the interdiffusion between PVDF and PMMA below the melting temperat... more In this study we investigated the interdiffusion between PVDF and PMMA below the melting temperature of PVDF by ATR-FTIR spectroscopy. The influence of the stereoregularity of different PMMA samples was studied. The PMMA tacticity showed a significant influence on the kinetics of the diffusion. Syndiotactic PMMA diffuses faster than isotactic and atactic PMMA which can be explained from the difference of chains stiffness between the two stereoisomers.
This paper presents results from a study of fibres extracted from Zostera marina eel-grass collec... more This paper presents results from a study of fibres extracted from Zostera marina eel-grass collected from the Baltic coast. This species of sea-grass is shown to contain small diameter (around 5 μm) fibres composed of ∼57% cellulose, ∼38% of non-cellulosic polysaccharides (mainly xylan) and ∼5% of residual matter so-called Klason lignin. This composition is quite different to that of commonly used terrestrial fibres. Single fibre stiffness values up to 28 GPa were measured. This stiffness combined with a low density could provide an attractive reinforcement for composite materials, and may be particularly suitable for use in bio-degradable structures.
The pH dependence of the tertiary structural changes and the quaternary reorganization of the αβ ... more The pH dependence of the tertiary structural changes and the quaternary reorganization of the αβ interface of oxyhemoglobin in solution has been previously observed in our laboratory. The present work aims to establish whether or not these progressive structural changes with pH result from proton exchange between the protein and the solvent. We therefore have used infrared spectroscopy, acid/base titration and 1H/2H exchange to assess the effect of external proton concentration on the structural and dynamic properties of the hemoglobin molecule in solution. This study is also performed on the carbonmonoxy form since the affinity of hemoglobin for CO is much higher than for oxygen. The present findings demonstrate that affinity changes are closely related to protein fluctuations, as well as structural modifications. An increased affinity, induced either by ligand replacement or pH variation, is associated with the constrained chains exhibiting a lower degree of fluctuation, together with a looser şβ interface association.
The environmentally sensitive fluorophore 2'-(N,N-dimethylamino)-6-naphthoyl-4-trans-cyclohexanoi... more The environmentally sensitive fluorophore 2'-(N,N-dimethylamino)-6-naphthoyl-4-trans-cyclohexanoic acid (DANCA) has been used to probe the apomyoglobin heme pocket. The unexpected polarity of this domain is generally interpreted as arising from dynamic dipolar relaxation of the peptide dipoles surrounding the heme pocket. In the present work we reexamine the photophysical properties of DANCA in a variety of solvents and complexed with apomyoglobin (apoMb) to further probe the heme pocket environment as a function of external solvent conditions. Absorption and excitation spectra in a number of solvents are consistent with the well-known pi*<--pi (LE) and pi*<--n (CT) electronic absorption transitions observed for naphthylamine derivatives. Dual emission is also a well-documented property of such derivatives. Based on the time scale of the heterogeneity in the decay of the DANCA fluorophore observed in a series of solvents, we propose that the emission properties of DANCA in apoMb are not uniquely attributable to dynamic relaxation events, but also reflect dual emission from both a long-lived, red CT state and the shorter-lived, blue LE state. The pH studies in the range of pH 5–9 of the emission properties of DANCA in apoMb support this hypothesis. They also suggest a specific interaction of DANCA with one or both of the pocket histidyl residues, which leads to a drastic static quenching and red shift of the bound DANCA fluorescence upon protonation. Similar effects are observed with increasing pressure, indicating that these two perturbations alter the DANCA-apoMb complex in a similar fashion. The pressure-induced form of the protein is distinct both energetically and structurally from the previously characterized acid intermediate, in that it is populated above pH 5 and retains a significant degree of integrity of the heme pocket.
Medium chain length polyhydroxyalkanoates (mcl-PHA) have been produced by fed-batch cultivation o... more Medium chain length polyhydroxyalkanoates (mcl-PHA) have been produced by fed-batch cultivation of Pseudomonas oleovorans. Using octanoic acid as the sole carbon substrate, a copolymer of 3-hydroxyoctanoate and 3-hydroxyhexanoate monomer units was accumulated up to 38 wt. % of cell mass. Mcl-PHA based nanocomposites have then been prepared using Cloisite 15A as nanofiller and characterized. Wide angle X-ray diffraction and rheological analysis of the nanocomposites highlighted their morphologies, which can be considered as well-intercalated structures. The thermal characteristics of mcl-PHA are not significantly affected by the nanofiller incorporation. On the contrary, the storage moduli of nanocomposites are greatly enhanced with increasing clay fraction due to the degree of dispersion of the nanoplatelets in the polymer matrix.
Medium chain length polyhydroxyalkanoates (mcl-PHA) have been produced by fed-batch cultivation o... more Medium chain length polyhydroxyalkanoates (mcl-PHA) have been produced by fed-batch cultivation of Pseudomonas oleovorans. Using octanoic acid as the sole carbon substrate, a copolymer of 3-hydroxyoctanoate and 3-hydroxyhexanoate monomer units was accumulated up to 38 wt. % of cell mass. Mcl-PHA based nanocomposites have then been prepared using Cloisite 15A as nanofiller and characterized. Wide angle X-ray diffraction and rheological analysis of the nanocomposites highlighted their morphologies, which can be considered as well-intercalated structures. The thermal characteristics of mcl-PHA are not significantly affected by the nanofiller incorporation. On the contrary, the storage moduli of nanocomposites are greatly enhanced with increasing clay fraction due to the degree of dispersion of the nanoplatelets in the polymer matrix.
The Journal of Steroid Biochemistry and Molecular Biology, 2015
Estrogens are steroid hormones that play a pivotal role in growth, differentiation and function o... more Estrogens are steroid hormones that play a pivotal role in growth, differentiation and function of reproductive and non-reproductive tissues, mediated through estrogen receptors (ERs). Estrogens are involved in different genomic and non-genomic cell signaling pathways which involve well-defined subcellular ER localizations. Thus, ER activity results from complex interplays between intrinsic binding properties and specific subcellular localization. Since these two factors are deeply intricate, we carried out, in a unique yeast cell context, a comparative study to better understand structure/function/ subcellular distribution relationships. This was carried out by comparing two ERs: the human ER a subtype (hERa) and the short form of the a isoform of the rainbow trout ER (rtERa S ). Their distinct binding properties to agonist and antagonist ligands and subcellular localizations were characterized in Saccharomyces cerevisiae yeast cells. An unexpected partial agonistic effect of ICI 182-780 was observed for rtERa S . Concomitant to distinct binding properties, distinct subcellular localizations were observed before and after ligand stimulation. Due to the unique cell context, the link between ERs intrinsic binding properties and subcellular localizations is partly unveiled and issues are hypothesized based on the role of cytoplasmic transient complexes which play a role in the ER cytoplasmic/nuclear partition, which in turn is critical for the recruitment of co-regulators in the nucleus.
A high-pressure Fourier-transform infrared technique was used to probe the evolution of 'H bonds ... more A high-pressure Fourier-transform infrared technique was used to probe the evolution of 'H bonds inside the helical segments of myoglobin in relation to p2H, Tris concentration in the medium and iron-ligand nature. The analysis was focused on changes in the conformation-sensitive amide-I' band, reflecting the peptide C = 0 group stretching vibrations coupled to the in-plane N-2H bending and C = N stretching modes. From data obtained under high pressure, the strength of 2H bonds, inside the a-helical segments of the protein at atmospheric pressure, is not simply a function of p2H and salt concentration. At low Tris concentration (50 mM), the strength of these 2H bonds increases with p'H, whereas for a higher Tris concentration (100 mM) this strength is lower at p2H 7 than at p'H 6.0 or 8.5. It is also observed that the azidometmyoglobin molecule exhibits tighter intrahelical interactions and lower sensitivity to pressure than aquametmyoglobin. Information is also presented regarding interhelical interactions in relation to the solvent.
Biochimica et Biophysica Acta (BBA) - Lipids and Lipid Metabolism, 1985
GalN induced a rapid inhibition of prelabelled triacylglycerol secretion by hepatocytes isolated ... more GalN induced a rapid inhibition of prelabelled triacylglycerol secretion by hepatocytes isolated from fed rats in which this secretion occurred to a larger extent than in hepatocytes isolated from fasted rats. The data reported here suggest that GalN induces a rise of triacylglycerol synthesis by inhibiting the palmitate oxidation pathway and a decrease of triacylglycerol secretion through an early derangement of the secretory pathway.
Chalcogenide glass optical fibers possess very low optical losses in the middle infrared range fr... more Chalcogenide glass optical fibers possess very low optical losses in the middle infrared range from 2 to 12 mm. They were used to implement remote infrared spectroscopy, known as Fiber Evanescent Wave Spectroscopy (FEWS). Due to their hydrophobic behavior, such sensor is especially suitable for application in biology and medicine where water is a nuisance to detect relevant information. Moreover,
Chalcogenide glasses are a matchless material as far as mid-infrared (IR) applications are concer... more Chalcogenide glasses are a matchless material as far as mid-infrared (IR) applications are concerned. They transmit light typically from 2 to 12 μm and even as far as 20 μm depending on their composition, and numerous glass compositions can be designed for optical fibers. One of the most promising applications of these fibers consists in implementing fiber evanescent wave spectroscopy, which enables detection of the mid-IR signature of most biomolecules. The principles of fiber evanescent wave spectroscopy are recalled together with the benefit of using selenide glass to carry out this spectroscopy. Then, two large-scale studies in recent years in medicine and food safety are exposed. To conclude, the future strategy is presented. It focuses on the development of rare earth-doped fibers used as mid-IR sources on one hand and tellurium-based glasses to shift the limit of detection toward longer wavelength on the other hand.
For decades, the origin of the concentric ring pattern of bacterial swarming colonies has puzzled... more For decades, the origin of the concentric ring pattern of bacterial swarming colonies has puzzled microbiologists. It was hypothesized that a periodic water activity variation originates a phase transition within the extracellular matrix water H bond network, which switches on and off the exopolysaccharide auto-organization. Both rheological and infrared spectroscopy measurements respectively performed at a molecular scale and on a currently migrating colony, have given a physical insight into the mechanisms which underlie the switch between swarming and consolidation phases. Thanks to in situ and real time infrared microspectroscopy, and thanks to the brilliance of the infrared beam at SOLEIL synchrotron, here we demonstrate that Proteus mirabilis swarming is triggered by a periodic variation of water activity at the colony edge. A dynamic behavior emerges from the global properties of the multicellular entity which relies on the ability of the bacterial cells to tune exoproduct synthesis in order to undergo sharp transitions at a given water activity threshold.
The pH dependence of the tertiary structural changes and the quaternary reorganization of the a j... more The pH dependence of the tertiary structural changes and the quaternary reorganization of the a j interface of oxyhemoglobin in solution has been previously observed in our laboratory. The present work aims to establish whether or not these progressive structural changes with pH result from proton exchange between the protein and the solvent. We therefore have used infrared spectroscopy, acidlbase titration and 'HI2H exchange to assess the effect of external proton concentration on the structural and dynamic properties of the hemoglobin molecule in solution. This study is also performed on the carbonmonoxy form since the affinity of hemoglobin for CO is much higher than for oxygen. The present findings demonstrate that affinity changes arc closely related to protein fluctuations, as well as structural modifications. An increased affinity, induced either by ligand replacement or pH variation, is associated with the constrained chains exhibiting a lower degree of fluctuation, together with a looser a/3 interface association It was recently shown that the tertiary and quaternary structures of oxyhemoglobin in solution evolve continuously with pH [l]. This gradual development was probed by observing the SH stretching vibration of two cysteinyl residues: aCysl04 (G11) and /3Cysll2 (G14). These two cysteine residues are known to be strongly H-bonded and very sensitive to slight conformational changes which are undetectable by other techniques [2, 31. The H-bond affects the distance between the sulfur and hydrogen atoms and, consequently, perturbs the SH vibration. The aCysl04 (G11) residue provides detailed information on the tertiary constraints of the G chain a-helices whereas the thiol group of pCysll2 (G14) is sensitive to quaternary structural changes at the tight ap interface Although the functional properties of hemoglobin are known to be pH dependent [5 -81, the changes in interactions inside the protein, induced by the surrounding solvent, are not yet clearly understood. While the affinity of hemoglobin for oxygen increases with pH, the molecular constraints at the x p interface decrease and the intrachain H-bonds of the a chain become stronger [I]. To investigate further the effect of pH on the molecular organization of hemoglobin, the Fouriertransform infrared study of the stretching vibrations of SH was extended to the carbonmonoxy hemoglobin derivative since the partition coefficient between O2 and CO is about 260 for the fully ligated hemoglobin . Furthermore, in order to obtain a more general understanding of the mechanisms correlating the proton concentration in the external solvent with the dynamic properties of these two hemoglobin forms (0, and CO), we have studied their acid/base titration behaviour and their 'H/2H exchange variations with pH. Indeed, many protons of the protein are exchangeable with those of the external solvent [9 -121. Besides, Lumry and Rosenberg
Fiber evanescent wave spectroscopy (FEWS) was used to classify various foodborne pathogens (Liste... more Fiber evanescent wave spectroscopy (FEWS) was used to classify various foodborne pathogens (Listeria monocytogenes, Staphylococcus aureus and Salmonella enteritidis) using chemometric methods such as principal component analysis (PCA) and logistic-partial least squares (logistic-PLS). Spectra were recorded thanks to an IR optical fiber made of chalcogenide glasses. A limited segregation between the healthy and non healthy food has been observed using the PCA method, whereas a clear differentiation between bacteria strains was obtained using the PLS regression. FEWS coupled to chemometric methods is a promising technique for the identification of foodborne pathogens as it could provide a rapid identification and classification of the bacteria.
Chalcogenide glass optical fibers possess very low optical losses in the middle infrared range fr... more Chalcogenide glass optical fibers possess very low optical losses in the middle infrared range from 2 to 12 mm. They were used to implement remote infrared spectroscopy, known as Fiber Evanescent Wave Spectroscopy (FEWS). Due to their hydrophobic behavior, such sensor is especially suitable for application in biology and medicine where water is a nuisance to detect relevant information. Moreover, the design of the sensor using tapered fibers enables to improve the signal to noise ratio. Then, once coupled with unsupervised analysis technique such as Principle Component Analysis (PCA), it has been shown that this tool is efficient to differentiate between obese and control mice by recording their serum FEWS spectra. The same method has been carried out to detect in situ the both phenotypes of a bacterial culture.
Owing to their low losses in the 2-12 micrometers region, the Te- As-Se glass fibers are used for... more Owing to their low losses in the 2-12 micrometers region, the Te- As-Se glass fibers are used for infrared light transportation as well as sensing element based on evanescent wave absorption mechanism. The efficiency of the system is improved by tapering the fiber diameter in the sensing zone. With this kind of sensor, infrared spectra of biological tissue have been obtained. Spectra of mouse liver have been especially recorded in order to detect spectral differences between the healthy and the tumoral states of mouse liver.
Fiber evanescent wave spectroscopy (FEWS) explores the mid-infrared domain, providing information... more Fiber evanescent wave spectroscopy (FEWS) explores the mid-infrared domain, providing information on functional chemical groups represented in the sample. Our goal is to evaluate whether spectral fingerprints obtained by FEWS might orientate clinical diagnosis. Serum samples from normal volunteers and from four groups of patients with metabolic abnormalities are analyzed by FEWS. These groups consist of iron overloaded genetic hemochromatosis (GH), iron depleted GH, cirrhosis, and dysmetabolic hepatosiderosis (DYSH). A partial least squares (PLS) logistic method is used in a training group to create a classification algorithm, thereafter applied to a test group. Patients with cirrhosis or DYSH, two groups exhibiting important metabolic disturbances, are clearly discriminated from control groups with AUROC values of 0.94+/-0.05 and 0.90+/-0.06, and sensibility/specificity of 86/84% and 87/87%, respectively. When pooling all groups, the PLS method contributes to discriminate controls, cirrhotic, and dysmetabolic patients. Our data demonstrate that metabolic profiling using infrared FEWS is a possible way to investigate metabolic alterations in patients.
The effects of pH on the a-subunit tertiary structure and on the alpI interface of the hemoglobin... more The effects of pH on the a-subunit tertiary structure and on the alpI interface of the hemoglobin molecule in aqueous solution are observed using Fourier transform infrared spectroscopy ofSH stretching vibrations from the a104 and 6112 cysteinyl residues.
Previously, we have identified an imperfect estrogen response element (rtERE) in the promoter of ... more Previously, we have identified an imperfect estrogen response element (rtERE) in the promoter of the rainbow trout vitellogenin gene. Although this ERE leads to a lower transcriptional activation, a better estradiol stimulation in vivo as compared to consensus ERE (EREcs) was observed. Here we examine the ability of recombinant human estrogen receptor R (rhERR) to bind DNA containing the EREcs or the natural imperfect rtERE, which contains three mismatches. At low salt concentration, whatever the ERE sequence, dissociation equilibrium constants of the specific rhERR-ERE complexes are similar (K D ) 2 nM) with the same stoichiometry. As salt concentration increases from 80 to 200 mM KCl, the affinity of the rhERR-rtERE complex largely diminishes whereas that of rhERR-EREcs seems less affected. Hence the nature of the interactions stabilizing these complexes is different: more ionic in rhERR-rtERE as compared to rhERR-EREcs. Moreover, kinetic measurements showed that specific rhERR-ERE complexes exhibit shorter half-lives (few seconds) and that the rhERR-EREcs complex is more stable (33 s) than the complex that formed with rtERE (19.8 s), in accordance with equilibrium binding results. Finally, dynamic studies of rhERR have shown that the protein fluctuations are damped when the salt concentration increases or when bound to ERE and all the more with rtERE. The interplay of affinity, complex half-lives, and protein dynamics in the transcriptional regulation of estrogen receptor is discussed.
In this study we investigated the interdiffusion between PVDF and PMMA below the melting temperat... more In this study we investigated the interdiffusion between PVDF and PMMA below the melting temperature of PVDF by ATR-FTIR spectroscopy. The influence of the stereoregularity of different PMMA samples was studied. The PMMA tacticity showed a significant influence on the kinetics of the diffusion. Syndiotactic PMMA diffuses faster than isotactic and atactic PMMA which can be explained from the difference of chains stiffness between the two stereoisomers.
This paper presents results from a study of fibres extracted from Zostera marina eel-grass collec... more This paper presents results from a study of fibres extracted from Zostera marina eel-grass collected from the Baltic coast. This species of sea-grass is shown to contain small diameter (around 5 μm) fibres composed of ∼57% cellulose, ∼38% of non-cellulosic polysaccharides (mainly xylan) and ∼5% of residual matter so-called Klason lignin. This composition is quite different to that of commonly used terrestrial fibres. Single fibre stiffness values up to 28 GPa were measured. This stiffness combined with a low density could provide an attractive reinforcement for composite materials, and may be particularly suitable for use in bio-degradable structures.
The pH dependence of the tertiary structural changes and the quaternary reorganization of the αβ ... more The pH dependence of the tertiary structural changes and the quaternary reorganization of the αβ interface of oxyhemoglobin in solution has been previously observed in our laboratory. The present work aims to establish whether or not these progressive structural changes with pH result from proton exchange between the protein and the solvent. We therefore have used infrared spectroscopy, acid/base titration and 1H/2H exchange to assess the effect of external proton concentration on the structural and dynamic properties of the hemoglobin molecule in solution. This study is also performed on the carbonmonoxy form since the affinity of hemoglobin for CO is much higher than for oxygen. The present findings demonstrate that affinity changes are closely related to protein fluctuations, as well as structural modifications. An increased affinity, induced either by ligand replacement or pH variation, is associated with the constrained chains exhibiting a lower degree of fluctuation, together with a looser şβ interface association.
The environmentally sensitive fluorophore 2'-(N,N-dimethylamino)-6-naphthoyl-4-trans-cyclohexanoi... more The environmentally sensitive fluorophore 2'-(N,N-dimethylamino)-6-naphthoyl-4-trans-cyclohexanoic acid (DANCA) has been used to probe the apomyoglobin heme pocket. The unexpected polarity of this domain is generally interpreted as arising from dynamic dipolar relaxation of the peptide dipoles surrounding the heme pocket. In the present work we reexamine the photophysical properties of DANCA in a variety of solvents and complexed with apomyoglobin (apoMb) to further probe the heme pocket environment as a function of external solvent conditions. Absorption and excitation spectra in a number of solvents are consistent with the well-known pi*<--pi (LE) and pi*<--n (CT) electronic absorption transitions observed for naphthylamine derivatives. Dual emission is also a well-documented property of such derivatives. Based on the time scale of the heterogeneity in the decay of the DANCA fluorophore observed in a series of solvents, we propose that the emission properties of DANCA in apoMb are not uniquely attributable to dynamic relaxation events, but also reflect dual emission from both a long-lived, red CT state and the shorter-lived, blue LE state. The pH studies in the range of pH 5–9 of the emission properties of DANCA in apoMb support this hypothesis. They also suggest a specific interaction of DANCA with one or both of the pocket histidyl residues, which leads to a drastic static quenching and red shift of the bound DANCA fluorescence upon protonation. Similar effects are observed with increasing pressure, indicating that these two perturbations alter the DANCA-apoMb complex in a similar fashion. The pressure-induced form of the protein is distinct both energetically and structurally from the previously characterized acid intermediate, in that it is populated above pH 5 and retains a significant degree of integrity of the heme pocket.
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Papers by Olivier Sire