TROPINONE REDUCTION IN ATROPA-BELLADONNA ROOT CULTURES

Transformed root cultures of Atropa belladonna accumulate the tropane alkaloid hyoscyamine and the precursors tropane-3 alpha-ol (tropine) and tropane-3 beta-ol (pseudotropine), depending on the culture stage. The latter are stereoisomeric reduction products of tropinone and are formed by two different NADPH-dependent tropinone reductases. Feeding experiments indicate that tropine and pseudotropine do not isomerize and that only tropine is incorporated into hyocyamine. The pseudotropine-forming tropinone reductase was purified, characterized and compared with other tropinone reductases reported so far. It has a molecular weight of 78 500 three identical subunits measured as 27 500 by SDS-PAGE, an IEP at pH 4.5, and a pH optimum at 6.25. The enzyme has a K-m of 90 mu M for tropinone and 21 mu M for NADPH. It catalyses the reduction of a limited number of analogues of tropinone, but with higher K-m values. None of the substrate analogues inhibited tropinone reduction. The regulatory role of the pseudotropine-forming tropinone reductase in tropane alkaloid metabolism is discussed.