Genetic analysis of a chromosomal region containing vanA and vanB, genes required for conversion of either ferulate or vanillate to protocatechuate in Acinetobacter

VanA and VanB form an oxygenative demethylase that converts vanillate to protocatechuate in microorganisms. Ferulate, an abundant phytochemical, had been shown to be metabolized through a vanillate intermediate in several Pseudomonas isolates, and biochemical evidence had indicated that vanillate also is an intermediate in ferulate catabolism by Acinetobacter. Genetic evidence supporting this conclusion was obtained by characterization of mutant Acinetobacter strains blocked in catabolism of both ferulate and vanillate, Cloned Acinetobacter vanA and vanB were shown to be members of a chromosomal segment remote from a supraoperonic cluster containing other genes required for completion of the catabolism of ferulate and its structural analogs, caffeate and coumarate, through protocatechuate. The nucleotide sequence of DNA containing vanA and vanB demonstrated the presence of genes that, on the basis of nucleotide sequence similarity, appeared to be associated with transport of aromatic compounds, metabolism of such compounds, or iron scavenging. Spontaneous deletion of 100 kb of DNA containing this segment does not impede the growth of cells with simple carbon sources other than vanillate or ferulate, Additional spontaneous mutations blocking vanA and vanB expression were shown to be mediated by IS1236, including insertion of the newly discovered composite transposon Tn5613. On the whole, vanA and vanB appear to be located within a nonessential genetic region that exhibits considerable genetic malleability in Acinetobacter. The overall organization of genes neighboring Acinetobacter vanA and vanB, including a putative transcriptional regulatory gene that is convergently transcribed and overlaps vanB, is conserved in Pseudomonas aeruginosa but has undergone radical rearrangement in other Pseudomonas species.