Busting the Unbreakable Lignin

New evidence shows anaerobic fungi can degrade the hardiest of plant components

The Science

There are three main ingredients to tackle when it comes to breaking down a plant for energy: Cellulose, hemicellulose and lignin, cumulatively called lignocellulose, are the primary components of hardy cell walls. It’s the cellulose and hemicellulose that eventually yield fuels, and removing lignin in order to do so is a resource-intensive process. Now, a new study shows how anaerobic fungi — the kind living in the stomachs of livestock — are actually capable of naturally modifying lignin. 

The Impact

Traditional methods for breaking down biomass use aerobic methods (those using oxygen) to remove lignin and transform the remaining components into biofuels and bioproducts. If anaerobic fungi could be used to effectively modify lignin without oxygen and make use of it, it would change the entire landscape of biomass production and bioproduct engineering.

Summary

That organisms make use of cellulose and hemicellulose is well-established: Across all life forms and with little exception, organisms have figured out how to create enzymes that break these components down into glucose. It’s that third component, lignin, that has historically been less of a resource to be extracted and more of an obstacle to breaking them down. 

Lignin is a structural component for the plant, like the brick-and-mortar of a building or the human skeleton. It’s more complex, and poisons enzymes and the microbes that make them. It’s known that aerobic fungi, like mushrooms, have some ability to slowly break down lignin (think of a fallen, decomposing tree in the forest). Because oxygen is essential to how these fungi break down lignin, oxygen has historically been presumed essential to any type of lignin breakdown — and lignin breakdown by anaerobic fungi therefore thought to be impossible. 

And yet, little chemical clues left some researchers wondering if perhaps the accepted narrative wasn’t quite accurate, and maybe anaerobic fungi living in soils or the guts of livestock were more capable than presumed. After all, they’re able to produce enzymes highly efficient at making use of those other two components, which requires getting through the lignin. 

Researchers led by Michelle O’Malley’s lab at the University of California at Santa Barbara recently published a paper in Nature Microbiology offering irrefutable evidence that lignin can, in fact, be deconstructed without oxygen. Lead author Tom Lankiewicz cultivated anaerobic fungi (Neocallimastix californiae) on sorghum, switchgrass and poplar, then monitored changes to the plants’ lignocellulose composition, as well as degradation products that accumulated as the fungus did its work. 

The UC Santa Barbara team worked with researchers at the U.S. Department of Energy Joint Genome Institute, a DOE Office of Science User Facility at Lawrence Berkeley National Laboratory, and two DOE Bioenergy Research Centers, the Joint BioEnergy Institute and the Great Lakes Bioenergy Research Center. O’Malley is also affiliated with JBEI.

The JGI data portals and pipelines helped identify potential genes responsible for lignin deconstruction. (The JGI also conducted RNA sequencing and alignment to the reference fungal genome. The study’s pivotal NMR (nuclear magnetic resonance) data, gathered using magnetic fields to diagnose electronic structure and functionality on the atomic level, was generated at JBEI and analyzed by the GLBRC. By providing snapshots of the types of chemical bonds in plant cell walls, it enabled researchers to identify specific bond-breakages related to lignin modification happening in the absence of oxygen.

These findings offer the first real irrefutable proof that lignin can be deconstructed without oxygen. Identifying the precise mechanisms by which these fungi break down lignin will require further research, including confirming the genes and the enzymes those genes produce. If confirmed, and if they prove capable of scaling for industry application, the work could eventually yield new biotechnological opportunities. 

Contacts

BER Contact
Ramana Madupu, Ph.D
Program Manager
Biological Systems Sciences Division
Office of Biological and Environmental Research
Office of Science
Department of Energy
Ramana.Madupu@science.doe.gov

PI Contact
Michelle O’Malley, Ph.D
Professor, Department of Chemical Engineering
Department of Chemical Engineering
University of California, Santa Barbara
momalley@ucsb.edu

JGI Contact
Igor Grigoriev
Fungal Genomics Program Lead
DOE Joint Genome Institute
ivgrigoriev@lbl.gov 

Funding

The authors acknowledge their funding sources: Office of Biological and Environmental Research of the U.S. Department of Energy grant DE-SC0020420 (MAO), Institute for Collaborative Biotechnologies grants W911NF-09–D-0001 and W911NF-19-2-0026 (MAO), US Army Research Office contract W911NF-19-1-0010 (MAO), National Science Foundation grant MCB-1553721 (MAO), Office of Biological and Environmental Research of the U.S. Department of Energy Joint BioEnergy Institute (JBEI, http://www.jbei.org) contract DE-AC02–05CH11231 (Lawrence Berkeley National Laboratory), and the Office of Biological and Environmental Research of the U.S. Department of Energy Great Lakes Bioenergy Research Center (GLBRC, https://www.glbrc.org/) contract DE-SC0018409 (University of Wisconsin and Michigan State University). The work (proposal: 10.46936/10.25585/60000889) conducted by the U.S. Department of Energy Joint Genome Institute (https://ror.org/04xm1d337), a DOE Office of Science User Facility, is supported by the Office of Science of the U.S. Department of Energy operated under Contract No. DE-AC02-05CH11231.

Publication

Lankiewicz, T.S., Choudhary, H., Gao, Y. et al. “Lignin deconstruction by anaerobic fungi.” Nature Microbiology (2023). DOI: 10.1038/s41564-023-01336-8

Relevant Links

• JGI Feature: JGI at 25: Following Fungi that Pry Apart Plant Polymers
• JGI Release: Fungal Enzymes Team Up to More Efficiently Break Down Cellulose
• PNNL Release: Biofuel Tech Straight from the Farm