Kinetics and substrate specificity of the glycanase activity associated with particles of Klebsiella bacteriophage No. 13

There is a glycanase activity associated with the particles of Klebsiella bacteriophage No. 13 which catalyzes hydrolysis of O-β-d-glucopyranosyl-(1 → 4)-β-d-mannopyranose linkages in Klebsiella serotype 13 capsular polysaccharide. The initial glycanase reaction velocity (at 37° and the optimum pH of 7–8) exhibits hyperbolical dependence on low concentrations of substrate, with K_m = 170 μm (in repeating units), and V_max = 140 nmol/min (glucose equivalents, for 10¹⁰ plaque-forming units). The reaction is inhibited by reaction products (type-13 penta- and decasaccharides, one and two repeating-units), and also by the substrate at concentrations above 1mm. Of seventy-four heterologous Klebsiella, and two other bacterial capsular heteropolysaccharides, as well as two substrate derivatives tested, only the Klebsiella serotype-2, -22, and -37 glycans were also depolymerized by the phage-13 enzyme. The reacting polysaccharides consist of repeating units, characteristically containing (a) chain 3eq,1eq → 4eq,1eq dihexopyranosyl sequences around the susceptible linkages, with equatorial hydroxyl and hydroxymethyl groups at C-2 and C′-5 next to them, and (b) branch carboxyl groups that may have to be located within a certain distance from these linkages.