Measurements of [θ]222 of E. coli phosphatase on heating from 20° to 90° and subsequent cooling to 20° shows a gradual increase in [θ]222 on heating, while cooling shows a symmetric transition centered at 45°. Reheating and cooling shows the same phenomenon. Enzyme heated and cooled once is fully active. The activity of the enzyme depends on its storage conditions (buffer and pH for example), but such changes are least to some extent reversible, especially by heating in different solvents. We conclude the enzyme exists in several forms which are in slow equilibrium with each other, so that the enzyme responds slowly when heated and hence is not at equilibrium during heating/cooling experiments.
Keywords: Alkaline phosphatase, thermal hysteresis, nonequilibrium, subunit association, circular dichroism (CD) profile, differential scanning calorimetry (DSC)
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