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Anaerobic Limonene Metabolism in a Methanogenic Enrichment Involves a Glycine Radical Enzyme

Lookup NU author(s): Dr Anastasia ResteuORCiD

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This work is licensed under a Creative Commons Attribution 4.0 International License (CC BY 4.0).

Abstract

Limonene is a natural monoterpene omnipresent in human environments. It enters wastewater and is also metabolised in methanogenic digesters. A stable limonene-degrading methanogenic enrichment culture was investigated by metagenomic, metatranscriptomic and metaproteomic data sets to characterise the microbial community and identify the limonene degradation pathway. Thirty-two metagenome-assembled genomes revealed a complex community of bacteria and methanogenic archaea dominated by Candidatus Velamenicoccus archaeovorus as the top predator, contributing two-thirds of the reads in the metagenome. The presence of several fermenting bacteria (Anaerolineaceae, Aminidesulfovibrio, Smithellaceae, Lentimicrobium) indicated the recycling of necromass in a microbial loop. Only one hydrocarbon-activating enzyme system was expressed, a member of the alkyl- and arylsuccinate synthase family which is a glycine radical enzyme that adds fumarate to hydrocarbons. The limonenylsuccinate synthase gene encodes a modified substrate binding pocket with two smaller amino acids, suggesting an adaptation for the larger structure of limonene. The limonenylsuccinate synthase operon and a ring cleavage operon, as well as genes for the final syntrophic fermentation to acetate, hydrogen and formate were encoded in a Syntrophobacteraceae genome. Almost all genes for this degradation pathway were highly transcribed and expressed, demonstrating a catalytic role for glycine radical enzymes in methanogenic systems degrading limonene

Publication metadata

Author(s): Lonsing A, Martens GA, Resteu A, Kizina J, Wilkie I, Bahr A, Harder J

Publication type: Article

Publication status: Published

Journal: Environmental Microbiology

Year: 2025

Volume: 27

Issue: 11

Print publication date: 01/11/2025

Online publication date: 03/11/2025

Acceptance date: 27/11/2025

Date deposited: 29/01/2026

ISSN (print): 1462-2912

ISSN (electronic): 1462-2920

Publisher: Wiley

URL: https://doi.org/10.1111/1462-2920.70192

DOI: 10.1111/1462-2920.70192

Data Access Statement: Proteomic data are available at PRIDE (PXD025008). NGS sequences and MAGs are deposited at NCBI.

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