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.