Hyaluronic acid (HA) is a natural polysaccharide present in the connective tissues of vertebrates, often used in the cosmetics and pharmaceutical industries. HA is a strongly hydrophilic macromolecule forming highly swollen random coils in aqueous solutions. Although some authors reported the secondary and tertiary structures of HA chain, others brought convincing evidence contradicting this hypothesis. This study aims at investigation of the stability and dynamics of the temporary duplex HA structures at different NaCl concentrations by molecular-dynamics (MD) simulations. The tendency to duplex formation grows with NaCl concentration reaching its maximum at 0.6 M. This profile is a result of two counteracting NaCl-concentration dependent phenomena, the growing electrostatic-repulsion screening on one side and the disturbance of hydrogen-bonds formation on the other side. Although the weak intermolecular attraction cannot lead to long-lived secondary and tertiary structures, it may influence the properties of large HA macromolecules and concentrated HA solutions.
- MeSH
- chlorid sodný * MeSH
- kyselina hyaluronová * chemie MeSH
- simulace molekulární dynamiky MeSH
- statická elektřina MeSH
- voda MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
In this work, low molecular weight (17 kDa) hyaluronan was modified by dodecanoyl substituents. The activation of dodecanoic acid was mediated by benzoyl chloride towards the preparation of a mixed anhydride, which reacts in a second step with HA in water mixed with an organic solvent. The effect of the cosolvent was studied and showed an even distribution of substituents and higher reaction rate in water: 1,4-dioxane compared to water:tert-butanol where substituents occupy adjacent positions. The chemical characterization of the prepared derivatives was elucidated by NMR, FTIR spectroscopy, thermal analyses, and gas chromatography, while the distribution of substituents was evaluated by enzymatic degradation. Molecular-dynamics simulations reveal opposite solvent separations around HA and dodecanoyl chains, that is stronger in water:tert-butanol solution. The resulting incompatibility of solvation-shells of the two entities repels the reaction intermediates from the HA chain and drives them towards the already bound substituents, explaining the observed differences in the distribution evenness. Thus, the influence of the solvent on the reaction selectivity is observed by shielding reactive sites around HA. Therefore, a control of the distribution of the substituents was obtained by defining the concentration of HA and used cosolvent.
High pressure methods have become a useful tool for studying protein structure and stability. Using them, various physico-chemical processes including protein unfolding, aggregation, oligomer dissociation or enzyme-activity decrease were studied on many different proteins. Oligomeric protein dissociation is a process that can perfectly utilize the potential of high-pressure techniques, as the high pressure shifts the equilibria to higher concentrations making them better observable by spectroscopic methods. This can be especially useful when the oligomeric form is highly stable at atmospheric pressure. These applications may be, however, hindered by less intensive experimental response as well as interference of the oligomerization equilibria with unfolding or aggregation of the subunits, but also by more complex theoretical description. In this study we develop mathematical models describing different kinds of oligomerization equilibria, both closed (equilibrium of monomer and the highest possible oligomer without any intermediates) and consecutive. Closed homooligomer equilibria are discussed for any oligomerization degree, while the more complex heterooligomer equilibria and the consecutive equilibria in both homo- and heterooligomers are taken into account only for dimers and trimers. In all the cases, fractions of all the relevant forms are evaluated as functions of pressure and concentration. Significant points (inflection points and extremes) of the resulting transition curves, that can be determined experimentally, are evaluated as functions of pressure and/or concentration. These functions can be further used in order to evaluate the thermodynamic parameters of the system, i.e. atmospheric-pressure equilibrium constants and volume changes of the individual steps of the oligomer-dissociation processes.
