Purification and properties of 1,2-dehydroreticuline reductase from Papaver somniferum seedlings
1992
De-Eknamkul, W. | Zenk, M.H.
1,2-Dehydroreticuline reductase, the NADPH-dependent enzyme which reduces stereospecifically 1,2-dehydroreticuline to (R)-reticuline has been discovered in seedlings of the opium poppy (Papaver somniferum). The enzyme has been purified to apparent electrophoretic homogeneity by ammonium sulphate precipitation and five subsequent column chromatography steps. The isolated enzyme is a single polypeptide with Mr 30 000 and has a pH optimum at 8.5 and a temperature optimum at 30 degrees. The apparent Km values for 1,2-dehydroreticuline and NADPH are 10 and 7 micromolar, respectively. The enzyme mediates the transfer of the pro-S-hydride of NADPH to C-1 of 1,2-dehydroreticuline with high substrate specificity; neither 1,2-dehydronorreticuline nor 1,2-dehydrococlaurine are utilized by the enzyme. The enzyme activity is inhibited by (S)- and (R)-reticuline with I50 values of 0.05 and 0.10 mM, respectively. The reductase is a cytosolic enzyme and present only in morphinan alkaloid-containing plants. This highly species-, substrate- and stereospecific enzyme catalyses the provision of (R)-reticuline for the formation of morphinan alkaloids that possess also (R)-configuration at that chiral centre.
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