PURIFICATION AND CHARACTERIZATION OF AN NAD+-LINKED FORMALDEHYDE DEHYDROGENASE FROM THE FACULTATIVE RUMP CYCLE METHYLOTROPH ARTHROBACTER-P1

When Arthrobacter P1 is grown on choline, betaine, dimethylglycine or sarcosine, an NAD+-dependent formaldehyde dehydrogenase is induced. This formaldehyde dehydrogenase has been purified using ammonium sulphate fractionation, anion exchange- and hydrophobic interaction chromatography. The molecular mass of the native enzyme was 115 kDa +/- 10 kDa. Gel electrophoresis in the presence of sodium dodecyl sulphate indicated that the molecular mass of the subunit was 56 k Da +/- 3 kDa, which is consistent with a dimeric enzyme structure. After ammonium sulphate fractionation the partially purified enzyme required the addition of a reducing reagent in the assay mixture for maximum activity. The enzyme was highly specific for its substrates and the K(m) values were 0.10 and 0. 80 mM for formaldehyde and NAD+, respectively. The enzyme was heat-stable at 50-degrees-C for at least 10 min and showed a broad pH optimum of 8.1 to 8.5. The addition of some metal-binding compounds and thiol reagents inhibited the enzyme activity.