Heterologous Expression and Characterization of a Novel Mesophilic Maltogenic α-Amylase AmyFlA from Flavobacterium sp. NAU1659

A novel amylase AmyFlA from Flavobacterium sp. NAU1659, AmyFlA, was cloned and expressed in Esherichia coli. Based on phylogenetic and functional analysis, it was identified as a novel member of the subfamily GH13_46, sharing high sequence identity. The protein was predicted to consist of 620 amino acids, with a putative signal peptide of 25 amino acids. The enzyme was able to hydrolyze soluble starch with a specific activity of 352.97 U/mg at 50 degrees C in 50 mM phosphate buffer (pH 6.0). The K-m and V-max values of AmyFlA were respectively 3.15 mg/ml and 566.36 mu mol center dot ml(-1 center dot)min(-1) under optimal conditions. Its activity towards starch was enhanced by 63% in the presence of 1 mM Ca2+, indicating that AmyFlA was a Ca2+-dependent amylase. Compared to the reported maltogenic amylases, AmyFlA produced a lower variety of intermediate oligosaccharides at the start of the reaction so that the product mixture contained a higher proportion of maltose. These results indicate that AmyFlA may be potential application value in the production of high-maltose syrup.