Characterisation and substrate binding modes of exopolygalacturonase PGQ1 from Saccharobesus litoralis

Among the enzymes required for the efficient utilisation of pectin is polygalacturonase. Saccharobesus litoralis harbours two polygalacturonases belonging to glycoside hydrolase family 28 (GH28). One of them, PGQ1, cleaved polygalacturonate exolytically at the non-reducing end into monomeric units. It was most active at 60 °C and pH 8, with K_m and k_cat values of 2.3 mg/ml and 6.4 s^-1 respectively. Its homology model of a right-handed parallel β-helix core consisted of Asp297 as the general acid and either Asp276 or Asp298 as the general base. By inferring the substrate binding modes at the -1 and +1 subsites from known crystal structures, a hexagalacturonate could be docked into the highly electropositive binding cleft. Interestingly, while no residues were present in the vicinity to make up the +2 and +4 subsites, Arg361 and Arg430 could readily bind to the carboxyl groups of the galacturonates at the +3 and +5 subsites respectively. Structural comparison suggested that this binding pattern with missing subsites might be unique to closely related exopolygalacturonases. As S. litoralis grew much more slowly on extracellular galacturonate due to the lack of a transporter for the monosaccharide, PGQ1 probably functioned in the periplasm to help degrade oligopectates completely.Communicated by Ramaswamy H. Sarma.