Methanocaldococcus jannaschii uses a modified mevalonate pathway for biosynthesis of isopentenyl diphosphate

Abstract

Archaea have been shown to produce isoprenoids from mevalonate; however, genome analysis has failed to identify several genes in the mevalonate pathway on the basis of sequence similarity. A predicted archaeal kinase, coded for by the MJ0044 gene, was associated with other mevalonate pathway genes in the archaea and was predicted to be the "missing" phosphomevalonate kinase. The MJ0044-derived protein was tested for phosphomevalonate kinase activity and was found not to catalyze this reaction. The MJ0044 gene product was found to phosphorylate isopentenyl phosphate, generating isopentenyl diphosphate. Unlike other known kinases associated with isoprene biosynthesis, Methanocaldococcus jannaschii isopentenyl phosphate kinase is predicted to be a member of the aspartokinase superfamily.

FIG. 1.

Chromosomal organization of genes coding for mevalonate pathway enzymes in representative archaea, including Methanothermobacter thermoautotrophicum, Archaeoglobus fulgidus, Pyrococcus furiosus, Methanosarcina mazei, Sulfolobus solfataricus, and Aeropyrum pernix. The corresponding genes from M. jannaschii are indicated for comparison but are not colocalized on the chromosome. Open reading frame lengths are not drawn to scale.

FIG. 2.

The mevalonate pathway. The proposed archaeal modifications of the mevalonate pathway, including the reaction catalyzed by IP kinase, are indicated in the box.

FIG. 3.

³¹P NMR of IP kinase reaction products. The control reaction (A) and the IP kinase reaction (0.28 μg enzyme) (B) each contained 9 mM IP, 9 mM ATP, and 9 mM MgCl₂ and were incubated at 55°C for 30 min. After the spectrum shown in panel B was recorded, the sample was spiked with 9 mM ADP and 9 mM Pi and the spectrum shown in panel C was recorded. The intensity of the monophosphate ester at δ 4.2 (AMP?) did not change throughout the experiment.