Mireles, R. | bioRxiv Science

bioRxiv2025

A synthetic bacterium that degrades and assimilates poly(ethylene terephthalate)

Polyethylene terephthalate (PET) is the fourth most commonly used plastic worldwide. Like all plastics, post-consumer PET is poorly managed and accumulates in the environment, posing significant ecological threats. After 70 years of accumulation, microorganisms capable of degrading and assimilating PET have been isolated, demonstrating that PET can be broken down and converted into valuable cellular biomass or metabolic products. These natural isolates, however, are poorly characterized and challenging to genetically manipulate, which limits their further optimization and applicability. Here, we engineer a well-established synthetic biology chassis for the biodegradation and assimilation of PET. We modified the bacterium Pseudomonas putida KT2440 to heterologously express an active PET-hydrolytic enzyme extracellularly and to metabolize PET biodegradation products. The resulting strain, named PETBuster, was capable of growing on PET as the sole carbon source on solid and liquid media. We achieved 91% PET degradation after 21 days of culture, with a doubling time of 3.6 days, under mesophilic conditions. In this way, we demonstrate that PET fermentation is feasible, opening the door to the production of valuable chemicals from waste.

synthetic biology↗

bioRxiv2023

Collaborative metabolic curation of an emerging model marine bacterium, Alteromonas macleodii ATCC 27126

Inferring the metabolic capabilities of an organism from its genome is a challenging process, relying on computationally-derived or manually curated metabolic networks. Manual curation can correct mistakes in the draft network and add missing reactions based on the literature, but requires significant expertise and is often the bottleneck for high-quality metabolic reconstructions. Here, we present a synopsis of a community curation workshop for the emerging model marine bacterium Alteromonas macleodii ATCC 27126 and its genome database in BioCyc, focusing on pathways for utilizing organic carbon and nitrogen sources. Due to the scarcity of biochemical information or gene knock-outs, the curation process relied primarily on published growth phenotypes and bioinformatic analyses, including comparisons with related Alteromonas strains. We report full pathways for the utilization of the algal polysaccharides alginate and pectin in contrast to inconclusive evidence for one carbon metabolism and mixed acid fermentation, in accordance with the lack of growth on methanol and formate. Pathways for amino acid degradation are ubiquitous across Alteromonas macleodii strains, yet enzymes in the pathways for the degradation of threonine, tryptophan and tyrosine were not identified. Nucleotide degradation pathways are also partial in ATCC 27126. We postulate that demonstrated growth on nitrate as sole N source proceeds via a nitrate reductase pathway that is a hybrid of known pathways. Our evidence highlights the value of joint and interactive curation efforts, but also shows major knowledge gaps regarding Alteromonas metabolism. The manually-curated metabolic reconstruction is available as a "Tier-2" database on BioCyc. ImportanceMetabolic reconstructions are vital for the systemic understanding of an organisms ecology. Here, we report the outcome of a collaborative, interactive curation workshop to build a curated "metabolic encyclopedia" for Alteromonas macleodii ATCC 27126, a marine heterotrophic bacterium with widespread occurrence. Curating pathways for polysaccharide degradation, one-carbon metabolism, and others closed major knowledge gaps, and identified further avenues of research. Our study highlights how the combination of bioinformatic, genomic and physiological evidence can be harvested into a detailed metabolic model, but also identifies challenges if little experimental data is available for support. Overall, we show how an interactive get-together by a diverse group of scientists can advance the ecological understanding of emerging model bacteria, with relevance for the entire scientific community.

microbiology↗