A plant host, Nicotiana benthamiana, enables the production and study of fungal lignin-degrading enzymes.

Lignin has significant potential as an abundant and renewable source for commodity chemicals yet remains vastly underutilized. Efforts towards engineering a biochemical route to the valorization of lignin are currently limited by the lack of a suitable heterologous host for the production of lignin-degrading enzymes. Here, we show that expression of fungal genes in Nicotiana benthamiana enables production of members from seven major classes of enzymes associated with lignin degradation (23 of 35 tested) in soluble form for direct use in lignin activity assays. We combinatorially characterized a subset of these enzymes in the context of model lignin dimer oxidation, revealing that fine-tuned coupling of peroxide-generators to peroxidases results in more extensive C-C bond cleavage compared to direct addition of peroxide. Comparison of peroxidase isoform activity revealed that the extent of C-C bond cleavage depends on peroxidase identity, suggesting that peroxidases are individually specialized in the context of lignin oxidation. We anticipate the use of N. benthamiana as a platform to rapidly produce a diverse array of fungal lignin-degrading enzymes will facilitate a better understanding of their concerted role in nature and unlock their potential for lignin valorization, including within the plant host itself.

Enhanced Bioavailability and Microbial Biodegradation of Polystyrene in an Enrichment Derived from the Gut Microbiome of Tenebrio molitor (Mealworm Larvae).

As the global threat of plastic pollution has grown in scale and urgency, so have efforts to find sustainable and efficient solutions. Research conducted over the past few years has identified gut environments within insect larvae, including Tenebrio molitor (yellow mealworms), as microenvironments uniquely suited to rapid plastic biodegradation. However, there is currently limited understanding of how the insect host and its gut microbiome collaborate to create an environment conducive to plastic biodegradation. In this work, we provide evidence that T. molitor secretes one or more emulsifying factor(s) (30-100 kDa) that mediate plastic bioavailability. We also demonstrate that the insect gut microbiome secretes factor(s) (<30 kDa) that enhance respiration on polystyrene (PS). We apply these insights to culture PS-fed gut microbiome enrichments, with elevated rates of respiration and degradation compared to the unenriched gut microbiome. Within the enrichment, we identified eight unique gut microorganisms associated with PS biodegradation including Citrobacter freundii, Serratia marcescens, and Klebsiella aerogenes. Our results demonstrate that both the mealworm itself and its gut microbiome contribute to accelerated plastic biodegradation. This work provides new insights into insect-mediated mechanisms of plastic degradation and potential strategies for cultivation of plastic-degrading microorganisms in future investigations and scale-up.