Characterizing the secondary hydration shell on hydrated myoglobin, hemoglobin, and lysozyme powders by its vitrification behavior on cooling and its calorimetric glass-->liquid transition and crystallization behavior on reheating.
For hydrated metmyoglobin, methemoglobin, and lysozyme powders, the freezable water fraction of between approximately 0.3-0.4 g water/g protein up to approximately 0.7-0.8 g water/g protein has been fully vitrified by cooling at rates up to approximately 1500 K min-1 and the influence of cooling rate characterized by x-ray diffractograms. This vitreous but freezable water fraction started to crystallize at approximately 210 K to cubic ice and at approximately 240 K to hexagonal ice. Measurements by differential scanning calorimetry have shown that this vitreous but freezable water fraction undergoes, on reheating at a rate of 30 K min-1, a glass-->liquid transition with an onset temperature of between approximately 164 and approximately 174 K, with a width of between approximately 9 and approximately 16 degrees and an increase in heat capacity of between approximately 20 and approximately 40 J K-1 (mol of freezable water)-1 but that the glass transition disappears upon crystallization of the freezable water. These calorimetric features are similar to those of water imbibed in the pores of a synthetic hydrogel but very different from those of glassy bulk water. The difference to glassy bulk water's properties is attributed to hydrophilic interaction and H-bonding of the macromolecules' segments with the freezable water fraction, which thereby becomes dynamically modified. Abrupt increase in minimal or critical cooling rate necessary for complete vitrification is observed at approximately 0.7-0.8 g water/g protein, which is attributed to an abrupt increase of water's mobility, and it is remarkably close to the threshold value of water's mobility on a hydrated protein reported by Kimmich et al. (1990, Biophys. J. 58:1183). The hydration level of approximately 0.7-0.8 g water/g protein is approximately that necessary for completing the secondary hydration shell.
Sartor
G
G
Institut für Allgemeine, Anorganische, und Theoretische Chemie, Universität Innsbruck, Austria.
Hallbrucker
A
A
Mayer
E
E
D002152:Calorimetry, Differential Scanning; D003460:Crystallization; D005615:Freezing; D006358:Hot Temperature; D006801:Humans; D006860:Hydrogen Bonding; D007700:Kinetics; D008706:Methemoglobin / Q000737:chemistry*; D008786:Metmyoglobin / Q000737:chemistry*; D009113:Muramidase / Q000737:chemistry*; D011102:Polyhydroxyethyl Methacrylate; D011487:Protein Conformation; D013816:Thermodynamics; D014867:Water; D014961:X-Ray Diffraction
D014867:Water; D008786:Metmyoglobin; D011102:Polyhydroxyethyl Methacrylate; D008706:Methemoglobin; D009113:Muramidase