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E Pauthe

Publications and source records attributed to E Pauthe.

7 recordsLinked to original sources
PubMed2003

Refolding of a high molecular weight protein: salt effect on collapse.

Small-angle neutron scattering experiments were performed on dilute solutions of a high molecular weight protein (fibronectin, M = 580 kg/mol) in four cases: native conditions; unfolded state obtained by a denaturing agent (urea); and two badly refolded (or collapsed) states obtained by progressive elimination of the denaturing agent in salt-containing or salt-free solutions. Our main result is concerned by the conformation of the protein as the attempt for refolding is driven with or without salt. In salt-containing solution, we observe unambiguously that the protein chain collapses at large length scales but still obeys to a Gaussian statistics at short length scales. In other words, the globule embodies a large quantity of solvent compared to the compact situation. In salt-free solutions, the badly refolded protein is not globular but displays both a coil-like and an open conformation at large length scales and a local high density area. This behavior is discussed with respect to the scaling theories for polymers and polyampholytes.

Computer Simulation↗
PubMed2002

Temperature-induced beta-aggregation of fibronectin in aqueous solution.

Fibronectin structural reorganization induced by temperature has been investigated by Fourier-transform infrared (FT-IR) spectroscopy and light-scattering experiments. At 20 degrees C, from resolution enhanced by FT-IR spectra, 43% of beta sheet, 31% of turn and 26% of unordered structures were estimated. Static and quasi-elastic light-scattering results do not change significantly between 20 and 34 degrees C. Just below 50 degrees C, a decrease of 1/3 of beta sheet structures contents is observed, concomitantly with a corresponding increase of turn. The contribution of disordered structures is found to be temperature-independent. Above 50 degrees C, our data reveals the formation of intermolecular hydrogen bonding leading to the formation of intermolecular beta sheet structures. The IR band absorption at 1618 cm(-1) increases strongly as a function of temperature. The scattered intensity increases and becomes strongly q(2)-dependent. The dynamic structure factor is not a single exponential decay and becomes strongly dependent on the scattering angle. These results demonstrate that aggregation occurs in fibronectin solution. When temperature decreases, this aggregation is found irreversible. Fibronectin aggregation is driven by the formation of intermolecular hydrogen bonds responsible for intermolecular beta sheet structures.

Fibronectins↗
PubMed2000

Statistical conformation of human plasma fibronectin.

Fibronectin is a multifunctional glycoprotein (molecular mass, M = 530 kg/mol) of the extra cellular matrix (ECM) having a major role in cell adhesion. In physiological conditions, the conformation of this protein still remains debated and controversial. Here, we present a set of results obtained by scattering experiments. In "native" conditions, the radius of gyration (R(g) = 15.3 +/- 0.3 nm) was determined by static light scattering as well as small-angle neutron scattering. The hydrodynamic radius (R(H) = 11.5 +/- 0.1 nm) was deduced from quasi-elastic light scattering measurements. These results imply a low internal concentration compared to that of usual globular proteins. This is also confirmed by the ratio R(H)/R(g) = 0. 75 +/- 0.02 consistent with a Gaussian chain, whereas R(H)/R(g) = 1. 3 for spherical shaped molecules. However, adding a denaturing agent (urea 8 M) increases R(g) by a factor 2. This means that fibronectin "native" chain is not either completely unfolded. The average shape of fibronectin conformation was also probed by small-angle neutron scattering performed for reverse scattering vector q(-)(1) smaller than R(g) (0.2 < q(-)(1) < 15 nm). The measured form factor is in complete agreement with the form factor of a random string of 56 beads of 5 nm diameter. It rules out the possibility of unfolded chain as well as globular structures. These results have structural and biological implications as far as ECM organization is concerned.

Fibronectins↗
PubMed1999

Structural studies by 1H NMR and molecular modeling of peptide substrates of thermolysin in relation with its proteasic activity in water and glycerol.

In an attempt to explain the relationship between conformations of peptide substrates of thermolysin in natural form and the experimental enzymatic cleavages, five peptides of various length were studied in two solvents H2O and glycerol, which may mimic the catalytic environmental conditions. As NMR failed to define sufficiently rough constraints to ensure a convergence of a refinement process for such short and flexible peptides, the conformational space was first searched using the MCMM method. The generated structures were then clustered in families using a 0.3A rmsd criterion and the derived structural characteristics were compared to the experimental NMR parameters. In a first approach, the NMR consistent conformations were compared with the structure of a thermolysin bound peptidic inhibitor ZG(P)LL to characterize the free-ligand predisposition to be cleaved. Further molecular dynamic calculations were performed at 300 K on the conformations corresponding to families in agreement with the ZG(P)LL structure in order to obtain information on their stability and on the trajectories of the torsion angles involved in the active site recognition. In conclusion, for four studied peptides, some conformations were found to be in agreement with 5 of the 8 cleavages experimentally observed.

Glycerol↗
PubMed1997

Additional data about thermolysin specificity in buffer- and glycerol-containing media.

Synthesis and use of various substrates permit an improved approach to thermolysin-peptide recognition and elucidation of several new criteria affecting enzyme specificity. Nature and position of the recognized residue, role of adjacent amino acids, lateral chain hydrophobicity, and volume and length of peptides were all considered. Hydrolysis reactions were also carried out in the presence of glycerol; the effect of microenvironment modifications was quantitative, for example in inducing variations in catalytic reaction rates, and also qualitative, such as in influencing affinity.

Buffers↗