Biosynthesis of Uridine Diphospho-N-acetylmuramic Acid

Uridine-5'-diphospho-N-acetyl-2-amino-2-deoxy-3-O-lactylglucose:NADP-oxidoreductase, an enzyme which catalyzes the reduction of UDP-GlcNAc-enolpyruvate to form uridine-5'-diphospho-N-acetylmuramic acid, was further purified to 1520-fold purification from Enterobacter cloacae NRC 492. Its molecular weight was estimated to be 35,500 by Sephadex gel filtration method. The reductase was greatly stimulated by a number of monovalent cations (T1+ g K+ g NH4+ g Rb+ >> Li+ ≃ Na+). The activation by K+ gave a normal hyperbolic saturation curve with a Ka of 5.8 x 10-3 m. Hill plots suggested a single binding site for K+ in association with each catalytic center. The activation by K+ was instantaneous and did not cause any shift in the pH optimum. The increase in K+ concentration from 2 mm to 20 mm markedly lowered the apparent Km value for UDP-GlcNAc-enolpyruvate. The value changed from 10.5 x 10-6 m to 6.5 x 10-6m. Effect on the Km value for NADPH was small. The data were consistent with the proposal that K+ might be acting as a bridge between UDP-GlcNAc-enolpyruvate and the enzyme.