Characterization of a siderophore from Agrobacterium fabrum C58

The majority of free iron in an aerobic environment forms insoluble iron(III) compounds.  This prevalence of iron limitation has put sufficient selective pressure on many bacteria to gain the ability to biosynthesize, secrete, and uptake complex iron-chelating molecules called siderophores.  We previously isolated a siderophore from Agrobacterium fabrum C58.  Bioinformatic analysis of the genes involved in siderophore production suggests this molecule is assembled by a hybrid nonribosomal peptide synthetase (NRPS) / polyketide synthase (PKS).  Each of these NRPSs and PKSs were overproduced in E. coli and purified.  The solubility and activity of all of the NRPS components required coproduction with Atu3678, a member of the MbtH-like protein superfamily.  Both acyltransferase domains of the PKS components recognized malonyl-CoA, while the initiating acyl-CoA ligase domain recognized both C6 and C10 fatty acid substrates. When combined with bioinformatic analyses, our data suggest the siderophore is a linear molecule containing an N-terminal fatty acid, a central thiazole ring, and a C-terminal bicyclic ring structure formed by the enzymatic modification of L-3,4-dihydroxyphenylalanine.  Finally, using the genetic sequences coding for these enzymes in bioinformatic searches, we identified related gene clusters in several cyanobacteria.  Using a combination of biochemistry, bioinformatics, and structural analysis we have begun to dissect the chemical structure of the siderophore produced by A. fabrum C58.  We propose the name fabrubactin for this siderophore.  The identification of the enzymology involved in the making of this unusual siderophore will provide new tools for the production of modified natural products with supplemented or enhanced functionality.