PURIFICATION AND CHARACTERIZATION OF SALUTARIDINE - NADPH 7-OXIDOREDUCTASE FROM PAPAVER-SOMNIFERUM

An enzyme which stereoselectively reduces the Papaver alkaloid, salutaridine, to (7S)-salutaridinol (=forward reaction) using NADPH as cosubstrate was discovered in Papaver somniferum cell cultures and differentiated plants. Using a seven-step procedure, the enzyme was purified from cell suspension cultures to apparent electrophoretic homogeneity. The isolated enzyme is a single polypeptide of M(r) 52+/-3 x 10(3) displaying an isoelectric point of 4.4. The physiological forward reaction has a pH optimum at 6.0-6.5, the reverse reaction at pH 9.0-9.5. The apparent K(m) values (forward reaction) for salutaridine and NADPH are 23 muM and 125 muM, respectively. The enzyme mediates the highly substrate-specific transfer of the pro-S hydride (B-type) of NADPH to C-7 of salutaridine. The reductase is a cytosolic enzyme. Screening of isoquinoline-containing plants and cell cultures demonstrated that the enzyme occurs only in P. somniferum and P. bracteatum. The reductive step catalysed by the oxidoreductase renders the salutaridinol molecule ready for the formation of the oxide bridge which characterizes morphine and related opium alkaloids.