Fig. 8
From: Discovery of processive catalysis by an exo-hydrolase with a pocket-shaped active site
The mechanism of substrate-product assisted processive catalysis by HvExoI. a Mechanism of Glc displacement with a disaccharide. After the disaccharide (empty blue and filled grey squares) bound in −1 and +1 subsites (step 1) is hydrolysed and an aglycon diffuses away (step 2), Glc (cyan square) remains non-covalently trapped and oscillates (double arrow) between the −1 and +1 subsites. Glc (yellow square) is consolidated to the −1 subsite after an incoming substrate binds (step 3), which later advances to the catalytic site. Glc (red square) modifies its binding patterns and exits (large arrow) via an autonomous and transient lateral cavity (cylinder in dashed lines) formed near the catalytic site (step 4); a next hydrolytic cycle can begin. b Mechanism of Glc displacement with a polysaccharide to facilitate substrate-product assisted processive catalysis. After non-reducing (blue square) and penultimate (filled square) residues bind in a productive mode at the −1 and +1 subsites (step 1), the non-reducing Glc residue (yellow square) is cleaved off, with the remainder of the substrate attached (step 2). Glc (red square) modifies its binding patterns and is released (large arrow) via a lateral cavity (cylinder in dashed lines, step 3). Here, the next hydrolytic cycle continues with the same polysaccharide, where the polysaccharide, short of one Glc (filled squares) advances into the catalytic site after uninterrupted binding. Non-reducing (blue) and hydrolysed (cyan-yellow-red in (a), yellow-red in (b)) Glc is shown. c Structural basis of substrate-product assisted processive catalysis. Left: The substrate (orange spheres) slides into the active site (protein surface: domain 1-light grey, domain 2-dark grey). After hydrolysis, the Glc product (yellow spheres) exits from the −1 subsite through the lateral cavity into the bulk solvent. Right: detail of the active site. Circle line with arrows indicates directionality during processive catalysis. The image was generated using the coordinates of HvExoI:G6SG-OMe and HvExoI:G2SG-OMe complexes (Fig. 3c–d; orange spheres), and the Glc displacement snapshots based on PELE calculations (Fig. 6e–h; yellow spheres)
