Biosynthesis of terpenoids

Abstract

The putative catalytic domain of an open reading frame from Plasmodium falciparum with similarity to the ispF gene of Escherichia coli specifying 2 C ‐methyl‐ d ‐erythritol 2,4‐cyclodiphosphate synthase was expressed in a recombinant E. coli strain. The recombinant protein was purified to homogeneity and was found to catalyze the formation of 2 C ‐methyl‐ d ‐erythritol 2,4‐cyclodiphosphate from 4‐diphosphocytidyl‐2 C ‐methyl‐ d ‐erythritol 2‐phosphate at a rate of 4.3 µmol·mg −1 ·min −1 . At lower rates, the recombinant protein catalyzes the formation of 2‐phospho‐2 C ‐methyl‐ d ‐erythritol 3,4‐cyclophosphate from 4‐diphosphocytidyl‐2 C ‐methyl‐ d ‐erythritol 2‐phosphate and the formation of 2 C ‐methyl‐ d ‐erythritol 3,4‐cyclophosphate from 4‐diphosphocytidyl‐2 C ‐methyl‐ d ‐erythritol. Divalent metal ions such as magnesium or manganese are required for catalytic activity. The enzyme has a pH optimum at pH 7.0. Recombinant expression of the full‐length open reading frame afforded insoluble protein that could not be folded in vitro . The enzyme is a potential target for antimalarial drugs directed at the nonmevalonate pathway of isoprenoid biosynthesis.