Abstract
Glucosaminyl phosphatidylglycerol (GlcN-PG) was first identified in bacteria in the 1960s and was recently reported in Pseudomonas aeruginosa. Despite the important implications in altering membrane charge (by the modification of anionic PG with cationic glucosamine), the biosynthesis and functions of GlcN-PG have remained uncharacterized. Using bioinformatic and lipidomic analysis, we identified a 3-gene operon, renamed as gpgSDF, that is responsible for the biosynthesis and potential transport of GlcN-PG in P. aeruginosa: gpgS encodes a novel glycotransferase that is responsible for the modification of phosphatidylglycerol (PG) with N-acetylglucosamine (GlcNAc) to produce GlcNAc-PG, and gpgD encodes a novel deacetylase that removes the acetyl group from GlcNAc-PG to produce GlcN-PG. The third gene in the operon, gpgF, is predicated to encode a flippase whose activity remains to be experimentally verified. As expected, the heterologous expression of the gpgSDF operon in Escherichia coli resulted in production of both GlcNAc-PG and GlcN-PG. The identification of the biosynthetic genes of GlcN-PG paves the way for the investigation of its biological and pathological functions, which has significant implications in our understanding of the unique membrane physiology, pathogenesis and antimicrobial resistance of P. aeruginosa.