Understanding enzymatic depolymerization of lignin
EMSL Project ID
49622
Abstract
Producing fuels and chemicals from lignocellulosic biomass is a promising technology to meet the nation's goal of a secure and sustainable energy future. Valorization of lignin is a key component of this effort, as it is typically 15-28% of the total dry weight of biomass. While it is a significant potential source of revenue, it is currently a waste stream that is usually burned to generate power. Economically-viable conversion of lignin to useful molecular building blocks has been elusive. Enzymes have potential as a low cost conversion strategy since they can be used at or near ambient temperature and pressure. However, a better understanding of the mechanisms of microbial ligninases is needed to increase the activity of enzyme cocktails to industrially-relevant levels, as has occurred with cellulases. Despite much effort since their discovery in the 1970s the activity of individual enzymes, or combinations of enzymes, on polymeric lignin is not understood. Microbial conversion strategies involve oxidative reactions, which are known to polymerize as well as depolymerize lignin. Understanding the factors that affect this balance is critical to developing an economically viable route to lignin-derived chemical products. While much prior work has focused on discovery of new enzymes and catalysts, much less work has emphasized understanding the physicochemical aspects that affect the polymerization/depolymerization equilibrium and the product distributions. Moreover, most past studies on the mechanisms of lignin breakdown have involved small molecule model compounds containing linkages found in lignin. Research conducted on model lignin substrates is not representative of native lignin, particularly with respect to repolymerization of reactive fragments. A major factor limiting progress on enzymatic depolymerization of polymeric lignin has been the lack of an assay for depolymerization of polymeric lignin that provides rapid exploration of the large parameter space. We have developed a new assay for lignin depolymerization based on films of insoluble lignin that enables simultaneous assaying of 78 independent reactions. Using this new approach we can now perform a systematic study of lignin degrading enzymes (e.g., laccases, peroxidases) and enzyme mixtures for a range of important reaction conditions. However, the assay reports only the amount of mass that is solubilized by the reaction. To understand reaction mechanisms, it is essential to have a detailed characterization of the product distribution and also chemical changes in the substrate. While SNL has the capability to analyze chemical changes in the substrate by FTIR and XPS, the MS expertise of EMSL is needed to characterize the mass distribution of solubilized products in the 300 ul reaction volumes at an estimated 30 ppm. If efficient enzymatic depolymerization of lignin and conversion to valuable products could be achieved at an industrially-relevant rate and scale, it would have a major impact on the economic viability of the lignocellulosic industry. While that goal is a long way off, the proposed work has the potential to yield major advances in understanding of ligninolytic cocktails. This is demonstrated by the discovery of an unanticipated maximum in lignin mass solubilized for a simple laccase-mediator system in preliminary work. Success in this project will allow us to determine the optimum concentration range for each enzyme and also the optimum mixture or sequence of enzymes to most effectively release desired soluble products from lignin.
Project Details
Project type
Exploratory Research
Start Date
2016-10-27
End Date
2017-09-30
Status
Closed
Released Data Link
Team
Principal Investigator
Co-Investigator(s)