Deciphering the properties and reaction mechanism of anhydromevalonate phosphate decarboxylase, a prenylated flavin mononucleotide‐dependent enzyme in the archaeal mevalonate pathway

In the archaeal mevalonate pathway, the prototype of all existing mevalonate pathways, a unique intermediate, trans-anhydromevalonate phosphate, is decarboxylated to form isopentenyl phosphate. The key reaction is catalyzed by a 3-octaprenyl-4-hydroxybenzoate carboxy-lyase (UbiD) family decarboxylase, anhydromevalonate phosphate decarboxylase (EC:4.1.1.126). The yet-to-be-identified properties of the archaea-specific enzyme, such as the requirement for prenylated flavin mononucleotide (prFMN) as a coenzyme, were elucidated using an enzyme derived from the hyperthermophilic archaeon Aeropyrum pernix. The coenzyme can be supplied to the decarboxylase from coexisting prFMN synthase, which anaerobically catalyzes the prenylation of reduced flavin mononucleotide and subsequent cyclization. Kinetic analysis of A. pernix anhydromevalonate phosphate decarboxylase supported its physiological role in catalyzing the decarboxylation step and progressing the archaeal mevalonate pathway, which is characterized by lower ATP consumption than other mevalonate pathways and is therefore considered promising for future metabolic engineering. However, nuclear magnetic resonance and liquid chromatography-mass spectrometry analyses showed that the enzyme could form non-negligible amounts of secondary products, probably because of the reactivity of the intermediate cycloaddition adduct between prFMN and the substrate. This study provides deeper insights into the reaction mechanism of UbiD family decarboxylases via 1,3-dipolar cycloaddition.