Bacillus licheniformistrehalose-6-phosphate hydrolase structures suggest keys to substrate specificity

Abstract

Trehalose-6-phosphate hydrolase (TreA) belongs to glycoside hydrolase family 13 (GH13) and catalyzes the hydrolysis of trehalose 6-phosphate (T6P) to yield glucose and glucose 6-phosphate. The products of this reaction can be further metabolized by the energy-generating glycolytic pathway. Here, crystal structures ofBacillus licheniformisTreA (BlTreA) and its R201Q mutant complexed withp-nitrophenyl-α-D-glucopyranoside (R201Q–pPNG) are presented at 2.0 and 2.05 Å resolution, respectively. The overall structure ofBlTreA is similar to those of other GH13 family enzymes. However, detailed structural comparisons revealed that the catalytic site ofBlTreA contains a long loop that adopts a different conformation from those of other GH13 family members. Unlike the homologous regions ofBacillus cereusoligo-1,6-glucosidase (BcOgl) andErwinia rhaponticiisomaltulose synthase (NX-5), the surface potential of theBlTreA active site exhibits a largely positive charge contributed by the four basic residues His281, His282, Lys284 and Lys292. Mutation of these residues resulted in significant decreases in the enzymatic activity ofBlTreA. Strikingly, the281HHLK284motif and Lys292 play critical roles in substrate discrimination byBlTreA.