Enzymatic cleavage of model lignin dimers depends on pH, enzyme, and bond type
Abstract Lignin is composed of phenylpropanoid monomers linked by ether and carbon-carbon bonds to form a complex heterogeneous structure. Bond-specific studies of lignin-modifying enzymes (LMEs; e.g., laccases and peroxidases) are limited by the polymerization of model lignin substrates and repolym...
Saved in:
| Main Authors: | , , , , , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Nature Portfolio
2025-03-01
|
| Series: | Scientific Reports |
| Subjects: | |
| Online Access: | https://doi.org/10.1038/s41598-025-88571-7 |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| _version_ | 1849326595925344256 |
|---|---|
| author | Jenny R. Onley Kshitiz Gupta Markus de Raad Benjamin P. Bowen Stephen Tan Sam Yoder Kenneth L. Sale Anup K. Singh Blake A. Simmons Paul D. Adams Trent R. Northen Kai Deng |
| author_facet | Jenny R. Onley Kshitiz Gupta Markus de Raad Benjamin P. Bowen Stephen Tan Sam Yoder Kenneth L. Sale Anup K. Singh Blake A. Simmons Paul D. Adams Trent R. Northen Kai Deng |
| author_sort | Jenny R. Onley |
| collection | DOAJ |
| description | Abstract Lignin is composed of phenylpropanoid monomers linked by ether and carbon-carbon bonds to form a complex heterogeneous structure. Bond-specific studies of lignin-modifying enzymes (LMEs; e.g., laccases and peroxidases) are limited by the polymerization of model lignin substrates and repolymerization of cleavage products. Here we present a high throughput platform to screen LME activities on four tagged model lignin compounds that represent the β-O-4’, β-β’, 5–5’, and 4-O-5’ linkages in lignin. We utilized nanostructure-initiator mass spectrometry (NIMS) and model lignin compounds with tags containing perfluorinated and cationic moieties, which effectively limit polymerization and condensation of the substrates and their degrading products. Sub-microliter sample droplets were printed on the NIMS chip with a novel robotics method. This rapid platform enabled characterization of LMEs across a range of pH 3–10 and relative quantification of modified (typically oxidized), cleaved, and polymerized products. All tested enzymes oxidized the four substrates and cleaved the β-O-4’ and β-β’ substrates to monomeric products. We discovered that the active pH range depended on both the substrate and the enzyme type. This has important applications for biomass conversion to biofuels and bioproducts, where the relative percentages of different bond types in lignin varies depending on feedstock and chemical pretreatment methods. |
| format | Article |
| id | doaj-art-80e663d7ee62490591a925d8f4e34f0b |
| institution | Kabale University |
| issn | 2045-2322 |
| language | English |
| publishDate | 2025-03-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Scientific Reports |
| spelling | doaj-art-80e663d7ee62490591a925d8f4e34f0b2025-08-20T03:48:06ZengNature PortfolioScientific Reports2045-23222025-03-0115111110.1038/s41598-025-88571-7Enzymatic cleavage of model lignin dimers depends on pH, enzyme, and bond typeJenny R. Onley0Kshitiz Gupta1Markus de Raad2Benjamin P. Bowen3Stephen Tan4Sam Yoder5Kenneth L. Sale6Anup K. Singh7Blake A. Simmons8Paul D. Adams9Trent R. Northen10Kai Deng11Technology Division, Joint BioEnergy InstituteTechnology Division, Joint BioEnergy InstituteEnvironmental Genomics and Systems Biology Division, Lawrence Berkeley National LaboratoryEnvironmental Genomics and Systems Biology Division, Lawrence Berkeley National LaboratoryTechnology Division, Joint BioEnergy InstituteTechnology Division, Joint BioEnergy InstituteBiosecurity and Bioassurance Department, Sandia National LaboratoriesTechnology Division, Joint BioEnergy InstituteBiological Systems and Engineering Division, Lawrence Berkeley National LaboratoryTechnology Division, Joint BioEnergy InstituteTechnology Division, Joint BioEnergy InstituteTechnology Division, Joint BioEnergy InstituteAbstract Lignin is composed of phenylpropanoid monomers linked by ether and carbon-carbon bonds to form a complex heterogeneous structure. Bond-specific studies of lignin-modifying enzymes (LMEs; e.g., laccases and peroxidases) are limited by the polymerization of model lignin substrates and repolymerization of cleavage products. Here we present a high throughput platform to screen LME activities on four tagged model lignin compounds that represent the β-O-4’, β-β’, 5–5’, and 4-O-5’ linkages in lignin. We utilized nanostructure-initiator mass spectrometry (NIMS) and model lignin compounds with tags containing perfluorinated and cationic moieties, which effectively limit polymerization and condensation of the substrates and their degrading products. Sub-microliter sample droplets were printed on the NIMS chip with a novel robotics method. This rapid platform enabled characterization of LMEs across a range of pH 3–10 and relative quantification of modified (typically oxidized), cleaved, and polymerized products. All tested enzymes oxidized the four substrates and cleaved the β-O-4’ and β-β’ substrates to monomeric products. We discovered that the active pH range depended on both the substrate and the enzyme type. This has important applications for biomass conversion to biofuels and bioproducts, where the relative percentages of different bond types in lignin varies depending on feedstock and chemical pretreatment methods.https://doi.org/10.1038/s41598-025-88571-7LigninLaccasePeroxidaseMass spectrometry |
| spellingShingle | Jenny R. Onley Kshitiz Gupta Markus de Raad Benjamin P. Bowen Stephen Tan Sam Yoder Kenneth L. Sale Anup K. Singh Blake A. Simmons Paul D. Adams Trent R. Northen Kai Deng Enzymatic cleavage of model lignin dimers depends on pH, enzyme, and bond type Scientific Reports Lignin Laccase Peroxidase Mass spectrometry |
| title | Enzymatic cleavage of model lignin dimers depends on pH, enzyme, and bond type |
| title_full | Enzymatic cleavage of model lignin dimers depends on pH, enzyme, and bond type |
| title_fullStr | Enzymatic cleavage of model lignin dimers depends on pH, enzyme, and bond type |
| title_full_unstemmed | Enzymatic cleavage of model lignin dimers depends on pH, enzyme, and bond type |
| title_short | Enzymatic cleavage of model lignin dimers depends on pH, enzyme, and bond type |
| title_sort | enzymatic cleavage of model lignin dimers depends on ph enzyme and bond type |
| topic | Lignin Laccase Peroxidase Mass spectrometry |
| url | https://doi.org/10.1038/s41598-025-88571-7 |
| work_keys_str_mv | AT jennyronley enzymaticcleavageofmodellignindimersdependsonphenzymeandbondtype AT kshitizgupta enzymaticcleavageofmodellignindimersdependsonphenzymeandbondtype AT markusderaad enzymaticcleavageofmodellignindimersdependsonphenzymeandbondtype AT benjaminpbowen enzymaticcleavageofmodellignindimersdependsonphenzymeandbondtype AT stephentan enzymaticcleavageofmodellignindimersdependsonphenzymeandbondtype AT samyoder enzymaticcleavageofmodellignindimersdependsonphenzymeandbondtype AT kennethlsale enzymaticcleavageofmodellignindimersdependsonphenzymeandbondtype AT anupksingh enzymaticcleavageofmodellignindimersdependsonphenzymeandbondtype AT blakeasimmons enzymaticcleavageofmodellignindimersdependsonphenzymeandbondtype AT pauldadams enzymaticcleavageofmodellignindimersdependsonphenzymeandbondtype AT trentrnorthen enzymaticcleavageofmodellignindimersdependsonphenzymeandbondtype AT kaideng enzymaticcleavageofmodellignindimersdependsonphenzymeandbondtype |