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Abstract: E. coli aspartate transcarbamylase (ATCase) is a 310 kDa allosteric enzyme which catalyses the first committed step in pyrimidine biosynthesis. The binding of its substrates, carbamylphosphate and aspartate, induces significant conformational changes. This enzyme shows homotropic cooperative interactions between the catalytic sites for the binding of aspartate. This property is explained by a quaternary structure transition from T state (aspartate low affinity) to R state (aspartate high affinity) accompanied by a 5% increase of radius of gyration of ATCase. The same quaternary structure change is observed upon binding of the bisubstrate analogue PALA (N-(phosphonacetyl)-l-aspartate. Owing to the large incoherent neutron scattering cross-section of the hydrogen atom and the abundance of this element in proteins, inelastic neutron scattering gives a global view of protein dynamics as sensed via the individual motions of its hydrogen atoms. We present neutron scattering results of the local dynamics (few angstroms), at short time (few tens of picoseconds), of ATCase in T and R forms. Compared to the T form, we observe an increased mobility of the protein in the R form that we associate to an increase of accessible surface area to the solvent. Beyond this specific result, this highlights the key role of the accessible surface area (ASA) in dynamic contribution to inelastic neutron data in the picosecond time scale. In particular, we want to stress out (i) that a difference at the picosecond time scale does not allow to conclude to a difference in the dynamics at a longer time scale and to address whether the T state is looser than the R state (ii) how challenging is, any comparison in terms of general dynamics (tense or relaxed) between dynamic values deduced from experimental neutron data on proteins with different sequences and therefore ASA. This caveat holds particularly when comparing dynamics of a mesophile with the corresponding extremophile. [Copyright &y& Elsevier]

Abstract: We have studied the influence of pressure on structure and dynamics of a small protein belonging to the enzymatic catalysis: the bovine pancreatic trypsin inhibitor (BPTI). Using a copper-beryllium high-pressure cell, we have performed small angle neutron scattering (SANS) experiment on NEAT spectrometer at HMI (Berlin, Germany). In the SANS configuration, the evolution of the radius of gyration and of the shape of the protein under pressures up to 6000 bar has been studied. When increasing pressure from atmospheric pressure up to 6000 bar, the pressure effects on the global structure of BPTI result on a reduction of the radius of gyration from 13.4 Å down to 12.0 Å. Between 5000 and 6000 bar, some transition already detected by FTIR [N. Takeda, K. Nakano, M. Kato, Y. Taniguchi, Biospectroscopy, 4, 1998, pp. 209–216] is observed. The pressure effect is not reversible because the initial value of the radius of gyration is not recovered after pressure release. By extending the range of wave-vectors to high q, we have observed a change of the form factor (shape) of the BPTI under pressure. At atmospheric pressure BPTI exhibits an ellipsoidal form factor that is characteristic of the native state. When the pressure is increased from atmospheric pressure up to 6000 bar, the protein keeps its ellipsoidal shape. The parameters of the ellipsoid vary and the transition detected between 5000 and 6000 bar in the form factor of BPTI is in agreement with the FTIR results. After pressure release, the form factor of BPTI is characteristic of an ellipsoid of revolution with a semi-axis a, slightly elongated with respect to that of the native one, indicating that the pressure-induced structural changes on the protein are not reversible. The global motions and the internal dynamics of BPTI protein have been investigated in the same pressure range by quasi-elastic neutron scattering experiments on IN5 time-of-flight spectrometer at ILL (Grenoble, France). The diffusion coefficients D and the internal relaxation times <τ 2> of BPTI deduced from the analysis of the intermediate scattering functions show a slowing down of protein dynamics when increasing pressure. [Copyright &y& Elsevier]

We propose a new method for the analysis of high-resolution incoherent quasi-elastic neutron scattering spectra of water in porous media and apply it, in particular, to a case of water in hydrated tricalcium silicate. We plot the normalized frequency-dependent susceptibility as a function of a scaling variable ω/ωp, where ωp is the peak position of the α-relaxation. In this way, we are able to scale QENS data from a range of Q into a single master curve. We then fit this master curve with an empirical formula for the susceptibility proposed by Bergman (J. Appl. Phys. 88 (2000) 1356) to obtain three independent parameters previously introduced by us to describe the dynamic state of hydration water in a curing cement paste. Specifically, we extract q, β, and τ¯ where, q is the fraction of glassy water, and β and τ¯ the two parameters associated with the α-relaxation of the glassy water. [Copyright &y& Elsevier]