Purification and biochemical characterization of stable alkaline protease Prot-2 from Botrytis cinerea

► An extracellular alkaline protease (Prot-2) selectively secreted by Botrytis cinerea growing in medium containing Spirulina algae as inducer was purified. ► Prot-2 has a monomeric structure, is active in the pH range 5.0-9.0 and shows an optimal temperature of activity at 50 °C. Prot-2 is thermostable, and activated by Ca2+. ► Disulfide played a role in enzyme activity and/or stability. ► Prot-2 showed extreme stability towards non-ionic surfactants (5% Tween 20, 5% Triton X-100 and Nonidet P-40). ► It was relatively stable in 25% aqueous/organic solvent mixtures and was activated by oxidizing agents (H2O2 and sodium perborate).