Engineering of anp efficient mutant of Neisseria polysaccharea amylosucrase for the synthesis of controlled size maltooligosaccharides.

摘要: Amylosucrase from Neisseria polysaccharea naturally catalyzes the synthesis of α-1,4 glucans sucrose. The product profile is quite polydisperse, ranging soluble chains called maltooligosaccharides to high-molecular weight insoluble amylose. This enzyme was recently subjected engineering its active site enable recognition non-natural acceptor substrates. Libraries variants were constructed and screened on sucrose, allowing identification a mutant that showed 6-fold enhanced activity toward sucrose compared wild-type enzyme. Furthermore, unprecedented, as only controlled size (2 < DP < 21) with narrow polydispersity observed. variant, containing 9 mutations in site, characterized at both biochemical structural levels. Its x-ray structure determined further investigated by molecular dynamics understand origins higher production small maltooligosaccharides, totally abolished glucan synthesis.