Discovery and engineering of bacterial lignin degrading enzymes

Lignin is a major component of woody biomass and the most abundant aromatic biopolymer on earth.  Valorization of this recalcitrant but rich source of high-value aryl compounds is fundamental to sustainable and carbon neutral bio-economy. A greener approach to meet this objective is the use of microbial enzymes as biocatalysts. Historically, fungal enzymes are considered the most efficient at lignin de-polymerization; however, recent advances in genomics have highlighted the potential of bacteria and their enzymes in lignin degradation – the focus of our research for lignin valorization. Using genomics and biochemical approaches, we have identified and characterized bacterial ligninases, such as, DypB - the first lignin degrading dye decolorizing peroxidase (DyP) from Rhodococcus jostii RHA1, and CopA, a multi-copper oxidase (MCO) from a coal bed metagenome. The enzyme-modified industrial lignins were characterized using 31P-NMR and GPC-MALS, while the production and identification of monoaryls (for e.g., vanillin, syringaldehyde and 2,6-dimethoxybenzo-quinone) was confirmed by HPLC and GC-MS. Structure-guided engineering was used to improve the catalytic efficiency of DypB and DyP2, a lignin-degrading DyP from Amycolatopsis sp 75iv2. Mechanistic insights, obtained from the structure-function studies, are being used to improving the catalytic efficiency of bacterial ligninases. Ongoing work includes the development of high-throughput screens to discover novel ligninases in metagenomic libraries prepared from diverse environments, as well as the subsequent characterization of these activities and using them to augment the bacterial deconstruction of biomass.