Biophysical, photochemical and biochemical characterization of a protease from Aspergillus tamarii URM4634

Circular dichroism (CD) and fluorescence spectroscopy (FS) were used to monitor the pH-dependent conformational and structural stability changes induced by temperature and UV light on the protease from Aspergillus tamarii URM4634 at different pH values. The formation of photoproducts, such as N-formylkynurenine, dityrosine and kynurenine, were monitored with FS. The pH-dependent melting temperatures (Tm) were determined using CD and FS from 20 to 90 °C. Conformational changes were correlated with the pH-dependent biochemical activities. CD revealed that the protease is rich in α-helices. Thermal denaturation was irreversible at all pH range and displayed Tm values from 42.8 to 67.8 °C (CD) and from 38 to 60.3 °C (FS), which the highest Tm was observed at pH 6. The light and temperature induced to the formation of photoproducts was more intense at high pH value. Despite the biochemical data shows optimum pH 9, the highest stability was at pH 6, maintaining 100% of activity after 24 h. The acquired data permits to select the best physicochemical parameters to secure the optimal activity and stability when used in biotechnological applications. Furthermore, the conformal changes induced by temperature in the protein are directly correlated with its level of biochemical activity.