Redefining the Role of the EryM Acetyltransferase in Natural Product Biosynthetic Pathways

Abstract

The GNAT (GCN5-related N-acetyltransferase) superfamily comprises enzymes with a conserved fold and diverse catalytic activities, including primarily acyl transfer, with a few examples of decarboxylation. EryM, a GNAT enzyme from Saccharopolyspora erythraea, has been implicated in both erythromycin and erythrochelin biosynthesis, with dual functionality as an acetyltransferase and a decarboxylase. Despite an historical association with malonyl-coenzyme A decarboxylation activity, the structural basis for this dual activity has remained unknown as its close homologs were identified with only acyl transfer activity. Here, crystal structures of EryM in its free form (2.5 [A]) and in complex with acetyl-CoA (2.9 [A]) reveal insights into the active site architecture and substrate interactions. Functional assays demonstrate that EryM catalyzes acyl transfer but lacks decarboxylation activity, challenging long-standing assumptions about its biosynthetic role. Comparative analysis of EryM and homologs in siderophore biosynthetic pathways highlights a conserved catalytic pocket with an essential His and identically positioned side chains common to GNAT enzymes for N-acyl transfer from CoA to primary hydroxylamine substrates. Bioinformatic analysis defines a large GNAT subfamily that is broadly distributed in the microbial world. These findings redefine EryM as an acetyltransferase and provide a foundation for understanding GNAT functional diversity in natural product biosynthesis.