Abstract
When D-glucosaminate dehydratase (GADH) was incubated with D-glucosaminate (GlcNA) in veronal buffer (VB ; 0. 01M, pH 8. 0), GlcNA was converted stoichiometrically to glyceraldehyde, pyruvate, and ammonia (aldolase reaction A). This reaction occurred in addition to the dehydratase reaction (conversion of GlcNA to 2-keto-3-deoxy-D-gluconate and ammonia : α, β-elimination reaction, B). The ratio of the activities (A : B) was about 1 : 4. However, in potassium phosphate buffer (KPB ; 0. 04M, pH 8. 0), the aldolase reaction was inhibited to 3-4% of that in VB, and also inhibited by various derivatives of glycerol, in particular, glycerol-3-phosphate (glycerol-3-P) and glyceraldehyde-3-phosphate (glyceraldehyde-3-P) in VB. The native enzyme was inhibited by incubation with 0. 1M EDTA, and the activity was restored by incubation of the EDTA-treated enzyme with (Mn2 + + pyridoxal 5'-phosphate (PLP)). When the EDTA-treated enzyme was incubated with (Mn2+ + PLP + glycerol-3-P), the activity of reaction B increased to 131% but that of reaction A decreased to 21%. These results suggested that Mn2+, PLP, and the phosphate group of glycero1-3-P are involved in formation of the active enzyme. In the case of the aldolase reaction, Mn2+ion, which might be essential for the reaction, is chelated by the phosphate group of glycerol-3-P with resultant inhibition of the aldolase reaction.
