α-Amylase From Penicillium brevicompactum: Enzymatic Properties and Saccharification

Although α-amylases are critical biocatalysts in starch processing, fungal α-amylase diversity remains largely untapped. This study presents the first systematic characterization of an α-amylase from Penicillium brevicompactum NRRL864. Maximum α-amylase activity of 195 U/mL was obtained at pH 6.0 and 55°C under optimized fermentation conditions. These conditions consisted of 0.75% starch, 3% peptone, 0.5% NaCl, incubation at 20°C for 7 days. A novel α-amylase gene (DF) was identified via phylogenetic analysis and heterologously expressed in Escherichia coli . The gene encodes a 551-amino-acid α-amylase with an open reading frame of 1656 bp, showing 68.36% sequence identity to the α-amylase from Penicillium expansum . The DF enzyme displayed a specific activity of 489.4 U/mg under optimal conditions of 50°C and pH 6.0, and a 124.2% activity enhancement with 1 mM Ca 2 ⁺, indicating Ca 2 ⁺-dependence. The activity of DF was inhibited by a range of chemical reagents, including EDTA and urea. Substrate specificity assays confirmed that the DF enzyme exhibited optimal activity in hydrolyzing soluble starch. DF-mediated saccharification of corn starch yielded a reducing sugar concentration of 317.9 µg/mL within 70 min, confirmed by thin-layer chromatography. Beyond identifying P. brevicompactum as a novel α-amylase source, this work demonstrates DF as a Ca 2 ⁺-responsive GH13 enzyme with significant potential for industrial applications in biofuel and sweetener production.