The widespread utility of isoprenoids has recently sparked interest in efficient synthesis of isoprene-diphosphate precursors. Current efforts have focused on evaluating two-step "isoprenol pathways," which phosphorylate prenyl alcohols using promiscuous kinases/phosphatases. The convergence on isopentenyl phosphate kinases (IPKs) in these schemes has prompted further speculation about the class's utility in synthesizing non-natural isoprenoids. However, the substrate promiscuity of IPKs in general has been largely unexplored. Towards this goal, we report the biochemical characterization of five novel IPKs from Archaea and the assessment of their substrate specificity using 58 alkyl-monophosphates. This study r... More
The widespread utility of isoprenoids has recently sparked interest in efficient synthesis of isoprene-diphosphate precursors. Current efforts have focused on evaluating two-step "isoprenol pathways," which phosphorylate prenyl alcohols using promiscuous kinases/phosphatases. The convergence on isopentenyl phosphate kinases (IPKs) in these schemes has prompted further speculation about the class's utility in synthesizing non-natural isoprenoids. However, the substrate promiscuity of IPKs in general has been largely unexplored. Towards this goal, we report the biochemical characterization of five novel IPKs from Archaea and the assessment of their substrate specificity using 58 alkyl-monophosphates. This study reveals the IPK-catalyzed synthesis of 38 alkyl-diphosphate analogs and discloses broad substrate specificity of IPKs. Further, to demonstrate the biocatalytic utility of IPK-generated alkyl-diphosphates, we also highlight the synthesis of alkyl-l-tryptophan derivatives using coupled IPK-prenyltransferase reactions. These results reveal IPK-catalyzed reactions are compatible with downstream isoprenoid enzymes and further support their development as biocatalytic tools for the synthesis of non-natural isoprenoids.
