Degradation of brown coal and 1-methylnaphthalene by a Micrococcus luteus isolate: Investigation of a novel aromatic degradation gene cluster containing paa genes

Aims To identify novel brown coal-degrading bacteria, and elucidate the biochemical pathways involved in brown coal degradation. Methods and results Four bacterial isolates were identified from the surface of Indonesian brown coal, which can utilize naphthalene and 1-methylnaphthalene as carbon sources for growth. The genome sequence of the best-performing Micrococcus luteus K1 strain was determined. A novel aromatic degradation gene cluster was identified, containing several paa genes normally involved in phenylacetic acid degradation, and also containing genes found on aromatic meta-cleavage pathways. 1-Naphthoic acid was generated from 1-methylnaphthalene by M. luteus K1 whole cell biotransformation, and was also utilized as a growth substrate by M. luteus K1. Recombinant ligase PaaK from the new gene cluster was shown to activate either phenylacetic acid or 1-naphthoic acid to their respective CoA esters, consistent with 1-naphthoyl CoA being an intermediate on the pathway. From metabolite analysis and annotation of the gene cluster, a new 1-methylnaphthalene degradation pathway was proposed, via a benzene oxide-oxepin ring opening. Recombinant mono-oxygenase and extradiol catechol dioxygenase enzymes from the gene cluster were expressed, showing activities consistent with the later steps of the proposed pathway. Conclusions A new M. luteus K1 isolate was identified as a brown coal degrader, whose genome contains an unusual aromatic degradation cluster containing paa genes. This cluster is hypothesized to be responsible for 1-methylnaphthalene degradation.