Fig. 4.

Picomolar levels of HEHEAA, a compound spontaneously formed from ethanolamine, induce gfp expression in Pseudomonas GM79 (pP_pipA-gfp). (A) HEHEAA formation from ethanolamine and HeGly (21–23). (B) Activation of gfp expression in the Pseudomonas GM79 (pP_pipA-gfp) bioassay by HEHEAA. (C) Influence of mutations in the genes flanking pipR on HEHEAA activation of P_pipA-gfp. The strains are AapB⁻, AapB transporter mutant; AapF, AapF PBP mutant; PipR⁻, PipR mutant; WT, wild type. Data are GFP fluorescence of cells grown with (blue bars) or without (white bars) 1 µM HEHEAA. B and C show mean RFU per OD₆₀₀ × 10 of four biological replicates, and the bars represent the SDs. (D) The MS2 fragmentation spectrum of active material purified from ethanolamine, [M+H]⁺ 163.1071 m/z, at 8.18 min (indicated by the diamond). (E) The MS2 fragmentation spectrum of chemically synthesized HEHEAA, [M+H]⁺ 163.1101 m/z, at 8.14 min.