Functional shifts of the dual-substrate phosphoribosyl isomerase PriA from primary to specialized metabolism in rare Actinomycetota

Evolutionary functional innovations can occur while gene families expand and two homologous (or analogous) genes co-occur. PriA is a dual-substrate enzyme family exclusive to Actinomycetota , with activities in l -histidine and l -tryptophan biosynthesis (HisA and TrpF activities) that evolved after the loss of the trpF gene. Since this gene loss, PriA has undergone multiple functional gain and loss events in central metabolism. Here, we report further evolutionary scenarios of PriA. First, in Ornithimicrobiaceae , a rare Actinomycetota family, PriA coexists with HisA, concomitant with the loss of its HisA but not TrpF activity, and is recruited in a physiological genome context. Second, a priA homologue, adeK , is located in the biosynthetic gene cluster of the specialized metabolites adechlorin and 2′-amino-2′-deoxyadenosine. The adeK gene is encoded in several genomes of the Streptosporangiaceae family and coexists with the expected and conserved priA . In this scenario, the PriA enzymes conserve both activities, whereas AdeK has only detectable levels of HisA activity, which might be related to specificity for a chlorinated intermediary during adechlorin biosynthesis. The adeK gene was first identified in strain ATCC-39365, previously designated as Actinomadura sp., but reclassified here as Nonomuraea sp. We further identified unprecedented adechlorin/2′-amino-2′-deoxyadenosine/pentostatin producers within the Microbispora and Streptosporangium genera. Phylogenomic analysis revealed additional recruitments of priA and hisG homologues into diverse uncharacterized biosynthetic gene clusters taxonomically related to the nucleoside antibiotics adechlorin, pentostatin and pyrazomycin. These findings highlight how enzyme family expansions and recruitment drive metabolic functional innovation in Actinomycetota , involving both gene loss and gain throughout central and specialized metabolism, and how evolutionary genome mining allows unbiased natural product discovery.