As attractive functional ingredients, maltooligosaccharides (MOS) are typically prepared by controlled enzymatic hydrolysis of starch. However, the random attack mode of amylase often leads to discrete product distribution, thereby reducing yields and purities. In this study, a novel glycoside hydrolase family 13 amylase AmyEs from marine myxobacteria Enhygromyxa salina was identified efficient maltohexaose (G6)-forming ability (40 %, w/ w). By deciphering external chain length, we found that the high density of alpha-1,6-branching points benefits the G6 formation of AmyEs with high purity (71-82 %), indicating the substrate selectivity of AmyEs toward high- branched starch. Based on this, asynchronous conversion strategy was designed to enhance specific MOS yield from corn starch by exploiting branching enzymes and AmyEs, and the purity and yield of G6 respectively increased by 9.5 % and 5 % compared to single AmyEs treatment. Our results demonstrate that combinatorial catalysis of MOS-forming amylases and branching enzymes provides a favorable industrial preparation of specific MOS.
