Catalytic Cycle of Glycoside Hydrolase BglX from Pseudomonas aeruginosa and Its Implication in Biofilm Formation.

BglX is a heretofore uncharacterized periplasmic glycoside hydrolase (GH) of the human pathogen Pseudomonas aeruginosa. X-ray analy-sis identifies it as a protein homodimer. The two active sites of the homodimer comprise catalytic residues provided by each monomer. This arrangement is seen in less than 2% of the hydrolases of known structure. In vitro substrate profiling shows BglX is a catalyst for β-(1→2) and β-(1→3) saccharide hydrolysis. Saccharides with β-(1→4) or β-(1→6) bonds, and the β-(1→4) muropeptides from the cell-wall pep-tidoglycan, are not substrates. Additional structural insights from X-ray analysis (including structures of a mutant enzyme-derived Mich-aelis complex, two transition-state mimetics, and two enzyme-product complexes) enabled the comprehensive description of BglX catalysis. The half-chair (4H3) conformation of the transition-state oxocarbenium species, the approach of the hydrolytic water molecule to the oxocarbenium species, and the stepwise release of the two reac-tion prod...