In Cinchona robusta (Rubiaceae) cell suspension cultures, the activity of the enzyme isopentenyldiphosphate isomerase (isopentenyl-POP isomerase) is transiently induced after addition of a homogenate of the phytopathogenic fungus Phytophthora cinnamomi. The enzyme catalyses the interconversion of isopentenyl-POP and dimethylallyl diphosphate (dimethylallyl-POP) and may be involved in the biosynthesis of anthrayuinone phytoalexins that accumulate rapidly after elicitation of Cinchona cells. From elicitor-treated C. robusta cells, two isoforms of isopentenyl-POP isomerase have been purified to apparent homogeneity in four chromatographic steps. The purified forms are monomeric enzymes of 34 kDa (isoform I) and 29 kDa (isoform 11), with K, values for isopentenyl-POP of 5.1 pM and 1.0 pM, respectively. Both isoforms require MnZ+ or Mg2+ as cofactor, isoform I1 showing a preference for Mn2+ with maximum activity at 1.5-2 mM. Isoform I was most active in the presence of 0.5-1.5 mM Mg" or in the presence of 0.5 mM MnZ+. A pH optimum of 7-7.8 was found for both forms and both were competitively inhibited by geranyl diphosphate (K, 96 pM for isoform I) and the transition state analogue 2-(dimethy1amino)ethyl diphosphate. Rechromatography of purified isoforms did not indicate any interconversion of both forms. Western blot analysis, using antibodies raised against isopentenyl-POP isomerase purified from Capsicum annuum, showed the presence of both isoforms in the crude protein extracts from C. robustu cells. Isoform I1 was specifically induced by elicitation, non-treated cells contained low activity of this isoform. The possible role of isopentenyl-POP isomerase in the biosynthesis of anthraquinones is discussed.