Abstract
Acyclic monoterpene primary alcohol: NADP^+ oxidoreductase, the key enzyme in the biosynthesis of monoterpene alcohols and iridoids in plants, is unstable and has been only poorly characterized. However we have established the conditions to stabilize the enzyme from Rauwolfia serpentina cells, followed by purification of it to homogeneity. It is a monomer with a molecular weight of about 44,000 and contains zinc ions. Various branched - chain allylic primary alcohols such as nerol (6), geraniol (7) and 10-hydroxygeraniol (1) were good substrates, but ethanol was inert. Menthol (16), cyclic monoterpene secondary alcohol, was also inert as a substrate, but inhibited the enzyme competitively. The enzyme exclusively requires NADP^+ or NADPH as the cofactor. Steady - state kinetic studies showed that the dehydrogenation proceeds by an ordered Bi - Bi mechanism. NADP^+ binds the enzyme first and then NADPH is the second product released from it. Neral, 10-hydroxygeranial (2) and 10-oxogeraniol (3) were enzymatically reduced in the presence of stereospecifically deuterium - labelled NADPH's. Capillary gas chromatography - mass spectrometric analysis of the products showed that the enzyme exclusively transfers the pro-R hydrogen at C-4 of NADPH.
