Biosynthesis of terpenoids: YchB protein of Escherichia coli phosphorylates the 2-hydroxy group of 4-diphosphocytidyl-2C-methyl-D-erythritol

A comparative analysis of all published complete genomes indicated that the putative orthologs of the unannotated ychB gene of Escherichia coli follow the distribution of the dxs, dxr, and ygbP genes, which have been shown to specify enzymes of the deoxyxylulose phosphate pathway of terpenoid biosyn thesis, thus suggesting that the hypothetical YchB protein also is involved in that pathway. To test this hypothesis, the E. coli ychB gene was expres sed in a homologous host. The recombinant protein was purified to homogenei ty and was shown to phosphorylate 4-diphosphocytidyl-2C-methyl-D-erythritol in an ATP-dependent reaction. The reaction product was identified as 4-dip hosphocytidyl-2C-methyl-D-erythritol 2-phosphate by NMR experiments with va rious C-13-labeled substrate samples. A C-14-labeled specimen of this compo und was converted efficiently into carotenoids by isolated chromoplasts of Capsicum annuum. The sequence of E. coli YchB protein is similar to that of the protein predicted by the tomato cDNA pTOM41 (30% identity), which had been implicated in the conversion of chloroplasts to chromoplasts.