Abstract
Thermotoga maritima
invertase (β-fructosidase), a member of the glycoside hydrolase family GH-32, readily releases β-
D
-fructose from sucrose, raffinose and fructan polymers such as inulin. These carbohydrates represent major carbon and energy sources for prokaryotes and eukaryotes. The invertase cleaves β-fructopyranosidic linkages by a double-displacement mechanism, which involves a nucleophilic aspartate and a catalytic glutamic acid acting as a general acid/base. The three-dimensional structure of invertase shows a bimodular enzyme with a five bladed β-propeller catalytic domain linked to a β-sandwich of unknown function. In the present study we report the crystal structure of the inactivated invertase in interaction with the natural substrate molecule α-
D
-galactopyranosyl-(1,6)-α-
D
-glucopyranosyl-β-
D
-fructofuranoside (raffinose) at 1.87 Å (1 Å=0.1 nm) resolution. The structural analysis of the complex reveals the presence of three binding-subsites, which explains why
T. maritima
invertase exhibits a higher affinity for raffinose than sucrose, but a lower catalytic efficiency with raffinose as substrate than with sucrose.