Crystallogenesis studies of proteins in agarose gel―mbined effect of high hydrostatic pressure and pH

摘要

The combined effect of both pressure and pH on the crystallization in agarose gel of thaumatin (0.1-230 MPa and pH 5.7-8.0) and the lysozymes from turkey (0.1-100 MPa and pH 4.0-6.0) and hen egg-white (0.1-75 MPa and pH 4.0-6.0) was investigated. For each condition, crystal morphology was examined and four crystal growth parameters determined (crystal number as a measure of nucleation, solubility, supersaturation and pressure-induced volume changes of crystallization). Comparison of data reveals a different crystallization behavior of thaumatin and the two lysozymes. First, thaumatin is more pressure tolerant in agarose gel than the lysozymes, with crystals growing up to 230 MPa for the former and only up to 75/100 MPa for the latter. Second, while an increase in pressure and pH increases nucleation of thaumatin crystals and decreases their solubility, one observes an inverse effect with the two lysozymes. Here, an increase in pressure but a decrease in pH increases nucleation and solubility. Third, pH influences the pressure-induced volume changes of crystallization, the strongest pH-dependence being found for turkey lysozyme. However, the volume changes are negative for thaumatin and positive for both lysozymes. These volume changes are correlated with solubility changes and are explained by pH- and pressure-induced alterations of the conformation of the three proteins, including their solvation shells.