Synthesis of New Hyperbranched α-Glucans from Sucrose by Lactobacillus reuteri 180 Glucansucrase Mutants

alpha-Glucans produced by glucansucrase enzymes of lactic acid bacteria attract strong attention as novel ingredients and functional biopolymers in the food industry. In the present study, alpha-helix 4 amino acid residues D1085, R1088, and N1089 of glucansucrase GTF180 of Lactobacillus reuteri 180 were targeted for mutagenesis both jointly and separately. Analysis of the mutational effects on enzyme function revealed that all D1085 and R1088 mutants catalyzed the synthesis of hyperbranched alpha-glucans with 15-22% branching (a1 -> 3,6) linkages, compared to 13% in the wild-type GTF180. In addition, besides native (a1 -> 6) and (a1 -> 3) linkages, all of the mutations introduced a small amount of (a1 -> 4) linkages (5% at most) in the polysaccharides produced. We conclude that alpha-helix 4 residues, especially D1085 and R1088, constituting part of the +2 acceptor binding subsite, are important determinants for the linkage specificity. The new hyperbranched alpha-glucans provide very interesting structural diversities and may find applications in the food industry.