Analysis and molecular cloning of genes involved in thiophene and furan oxidation by E. coli

Escherichia coli NAR30 can oxidize thiophenes, furans, and various other aromatic compounds as a result of successive mutations in three novel genes (thdA, C, and D). Some of the mutant genes involved in thiophene degradation have been cloned into pUC19. One of the resulting plasmids, pKA10, which carries a 3.8-kb chromosomal fragment from NAR30, conferred the ability to partially degrade furfuryl alcohol and thiophene methanol on a fadR atoC host strain. The location of the thd gene(s) on the plasmid pKA10 was found by insertion of a kanamycin resistance cassette. One such insertion into a BglII site inactivated the thd gene(s). Disappearance of two proteins correlated with loss of oxidation ability vs thiophenes and furans. Strains carrying the thdA mutation oxidize these substrates by an oxidase that can be coupled to methylene blue. However, degradation of thiophenes by NAR30 is incomplete, with no organic sulfur being released.