Calorimetric Investigation of Copper Binding in the N-Terminal Region of the Prion Protein at Low Copper Loading: Evidence for an Entropically Favorable First Binding Event

Although the Cu²⁺-binding sites of the prion protein have been well studied when the protein is fully saturated by Cu²⁺, the Cu²⁺-loading mechanism is just beginning to come into view. Because the Cu²⁺-binding modes at low and intermediate Cu²⁺ occupancy necessarily represent the highest-affinity binding modes, these are very likely populated under physiological conditions, and it is thus essential to characterize them in order to understand better the biological function of copper–prion interactions. Besides binding-affinity data, almost no other thermodynamic parameters (e.g., Δ<i>H</i> and Δ<i>S</i>) have been measured, thus leaving undetermined the enthalpic and entropic factors that govern the free energy of Cu²⁺ binding to the prion protein. In this study, isothermal titration calorimetry (ITC) was used to quantify the thermodynamic parameters (<i>K</i>, Δ<i>G</i>, Δ<i>H</i>, and <i>T</i>Δ<i>S</i>) of Cu²⁺ binding to a peptide, PrP­(23–28, 57–98), that encompasses the majority of the residues implicated in Cu²⁺ binding by full-length PrP. Use of the buffer <i>N</i>-(2-acetomido)-aminoethanesulfonic acid (ACES), which is also a well-characterized Cu²⁺ chelator, allowed for the isolation of the two highest affinity binding events. Circular dichroism spectroscopy was used to characterize the different binding modes as a function of added Cu²⁺. The <i>K</i>_d values determined by ITC, 7 and 380 nM, are well in line with those reported by others. The first binding event benefits significantly from a positive entropy, whereas the second binding event is enthalpically driven. The thermodynamic values associated with Cu²⁺ binding by the Aβ peptide, which is implicated in Alzheimer’s disease, bear striking parallels to those found here for the prion protein

Spectroscopic investigation of metal binding to the prion protein

EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Thermal properties by design : synthesis, thermal and mechanical analysis, and molecular modelling of novel cyanate ester resins

EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Synthesis, Thermal and Mechanical Analysis and Molecular Modelling of Novel Cyanate Ester Resins.

This work is concerned with the preparation of six dicyanate ester monomers, four of which are novel and the same four have been cured. The first dicyanate contains a arylene ether sulphone backbone, the second is two dicyanate of siloxane type backbone and the final three are dicyanates containing alkylene ether backbone of varying lengths where n = 1, 2, 3. All the dicyanates were prepared in good yield and purity. Characterisation of the monomers was undertaken using spectroscopic and chromatographic methods and elemental analysis. Thermal analysis was carried out on the cured arylene ether sulphone dicyanate and alkylene ether dicyanates using differential scanning calorimetry with varying heating rates. The monomers were blended with AroCy B10 and both the homopolymers and blends were analysed using thermogravimetric analysis and the effects of the blend stoichiometry are assessed and comparisons made with the homopolymers. Dynamic mechanical thermal analysis was employed to examine the values of glass transition temperature and tensile moduli. This was performed on the arylene ether sulphone dicyanate homopolymer and their blends with AroCy B10 and in the form of bending moduli on the alkylene ether dicyanate homopolymers and blends with AroCy B10. Thermomechanical analysis was carried out on both homopolymers and blends with AroCy B10, to examine the effects the backbone types have on the thermal expansion behaviour. Arrhenius kinetic parameters are presented for the catalysed thermal polymerisation and degradation reactions of the arylene ether sulphone dicyanate. An improved method for obtaining these parameters, using numerical methods to simulate the exothermic DSC curve produced during polymerisation, is compared to that of the traditional method. A parameter set (RDA-DR2. 21_Inv), previously optimised for use with cyanates, was employed to reproduce both the geometries and the physical and mechanical properties of poly(bis-4-(4-cyanatophenoxy)phenyl sulphone). Molecular dynamics simulations, carried out on the polymer structure have yielded a glass transition temperature (Tg) for the polycyanurates of between 237 - 247 C, with a density of between 1.31 -1.34 gcm-3 at 300 K. The same dicyanate was prepared and characterised using dynamic mechanical analysis and thermo-mechanical analysis. The former yields an empirical Tg of 246C and modulus of 1.57 GPa (at 150C), while the latter produces a CTE of 39 ppm/C (40-200C). The density obtained from the prepared polymer, recorded at 298 K, gave a value of 1.34gcm-3 . This value is in good agreement with that obtained from improved simulation data