Purification and characterization of cutinase from Botrytis cinerea and effect on blueberry cuticle

Abstract

An extracellular cutinase in Botrytis cinerea was isolated and its biochemical properties and effects on the penetration of pathogens were evaluated. This cutinase had a specific activity of 6.03 U/μg of protein after 22.33-fold purification, and yielded a recovery of 50.5%. The optimum pH and temperature for the enzymatic activity of cutinase were 9 and 45 °C, respectively. The cutinase was stable between 30 and 80 °C and pH 4.0–10.0. Fe3+ and Cu2+ strongly increased cutinase activity while Mn2+ completely inhibited it. The enzyme could maintain its activity in the presence of methanol, ethanol and isopropanol, but was significantly inhibited in acetonitrile, n-hexane and acetone. Kinetic parameters KM, Vmax and kcat were 0.21 mM, 303 μmol•min−1•mg−1 protein and 303.03 s−1 respectively. The purified cutinase could also destroy blueberry wax crystal and decrease the cuticle, indicating a positive role on the penetration of pathogens for blueberry fruits. This study provides useful information for fruit postharvest preservation.

Keywords

• Cutinase
• Purification
• Biochemical characterization
• Penetration of pathogens
• Cuticle