| The biosynthetic pathway of phenazine antibiotics produced in two separate bacterial strains has been studied. In Streptomyces antibioticus TU 2706, the biosynthesis of the monomeric saphenyl metabolites, and their dimerization products the esmeraldins, proceeds from phenazine-1,6-dicarboxylic acid (PDC) by chain extension with C-2 of acetate to 6-acetylphenazine-1-carboxylic acid, which is reduced to saphenic acid and incorporated into both halves of the esmeraldins. By feeding of chiral acetate and degradation and configurational analysis of the resulting saphenyl metabolites and esmeraldins, the chain extension process was found to proceed with overall inversion of configuration at the methyl group; this suggests that an intermediate β-keto acid is decarboxylated with inversion stereochemistry. Stable isotope-labeled chorismic acid, 2-amino-2-deoxyisochorismic acid (ADIC) and anthranilic acid failed to incorporate into the S. antibioticus phenazines, although the negative results from one or more of these feeding experiments might be due to cell wall impermeability to the administered putative substrate. The pathway of phenazine 1-carboxylic acid (PCA) biosynthesis in Pseudomonas fluorescens 2-79 was studied through incubations of cell-free extracts of E. coli expressing cassettes of the entire PCA biosynthetic genes cluster from P. fluorescens, or subsets of these genes, with various substrates. These incubation experiments allowed for the identification of ADIC, but not anthranilic acid, and trans -2,3-dihydro-2,3-dihydroxybenzoic acid (DHHA) as key intermediates in PCA biosynthesis. PDC was ruled out as a free intermediate in PCA formation. |
