Purification and Characterization of d-Glucosaminate Dehydratase from Pseudomonas fluorescens

D-Glucosaminate Aldolase Activity ofD-Glucosaminate Dehydratase fromPseudomonas fluorescensand Its Requirement for Mn2+Ion

“…4) In this work, we show that GADH also has PLP-dependent GlcNA aldolase activity, and that Mn 2 + ion is essential for the aldolase reation. In a preliminary report, we showed that the enzyme from P. fluorescens is a bifunctional enzyme.…”

Contribution to Catalysis and to Stability of the Essential Cysteine Residue at the Active Site ofd-Glucosaminate Dehydratase fromPseudomonas fluorescens

“…5) The purified enzyme (specific activity; about 600 units/mg protein) has about 14.5% to 90% holo activity in each preparation (the activity ratio in the absence and in the presence of PLP). The enzyme with low holo activity was used as the apoenzyme without any further treatment.…”

“…After incubation at 37°C for 15 min, the amount of KDGA formed was measured as described previously by monitoring the absorbance at 250 nm of the semicarbazone. 5 ) One unit of enzyme was defined as the amount of enzyme that catalyzed the formation of I tJmol of KDGA per min under the given conditions. Protein was measured by the Lowry method 6 ) with bovine serum albumin as the standard.…”

Mn2+inD-Glucosaminate Dehydratase fromPseudomonas fluorescens

“…Thus, incubation of the EDT A-treated enzyme at 37°C for 30 min with PLP, which may be removed from the enzyme during the treatment with EDTA, and Mn 2 + is nece~say to restore the maximum activity of enzyme. (4) no addition at 37°C for 30min, and D-GlcNA (20mM) was added to the reaction mixture, then enzyme activity was measured as described in Materials and Methods. a Proposed structures are shown in Fig.…”