Effect of reaction conditions on the specific activity of wild type Wzb. A. Effect of temperature. Endpoint reactions were carried out in 50 mM MES – 50 mM HEPES buffer, pH 8, containing 1% BSA in the presence of Cu2+, Co2+, Fe3+ (0.1 mM each) and Mn2+, Mg2+ (0.5 mM each). A 30-min pre-incubation was carried out at the desired temperature before addition of 2.5 mM p NPP. Reactions were then stopped after 10 min incubation with 100 mM (final) sodium vanadate (Na3VO4). B. Effect of pH. Endpoint reactions were carried out in 100 mM MES – 100 mM HEPES buffer with 1% BSA in the presence of Cu2+, Co2+, Fe3+ (0.1 mM each) and Mg2+, Mn2+ (at 0.5 mM each). Incubation for 30 min at 75°C was carried out before the addition of 2.5 mM p NPP. Reactions were stopped after 10 min. with 100 mM (final concentration) sodium vanadate.