Dataset Information

High-Resolution Screening for Marine Prokaryotes and Eukaryotes With Selective Preference for Polyethylene and Polyethylene Terephthalate Surfaces.

ABSTRACT: Marine plastic debris serve as substrates for the colonization of a variety of prokaryote and eukaryote organisms. Of particular interest are the microorganisms that have adapted to thrive on plastic as they may contain genes, enzymes or pathways involved in the adhesion or metabolism of plastics. We implemented DNA metabarcoding with nanopore MinION sequencing to compare the 1-month-old biomes of hydrolyzable (polyethylene terephthalate) and non-hydrolyzable (polyethylene) plastics surfaces vs. those of glass and the surrounding water in a Mediterranean Sea marina. We sequenced longer 16S rRNA, 18S rRNA, and ITS barcode loci for a more comprehensive taxonomic profiling of the bacterial, protist, and fungal communities, respectively. Long read sequencing enabled high-resolution mapping to genera and species. Using previously established methods we performed differential abundance screening and identified 30 bacteria and five eukaryotic species, that were differentially abundant on plastic compared to glass. This approach will allow future studies to characterize the plastisphere communities and to screen for microorganisms with a plastic-metabolism potential.

SUBMITTER: Marsay KS

PROVIDER: S-EPMC9042255 | biostudies-literature | 2022

REPOSITORIES: biostudies-literature

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Publications

High-Resolution Screening for Marine Prokaryotes and Eukaryotes With Selective Preference for Polyethylene and Polyethylene Terephthalate Surfaces.

Marsay Katherine S KS Koucherov Yuri Y Davidov Keren K Iankelevich-Kounio Evgenia E Itzahri Sheli S Salmon-Divon Mali M Oren Matan M

Frontiers in microbiology 20220412

Marine plastic debris serve as substrates for the colonization of a variety of prokaryote and eukaryote organisms. Of particular interest are the microorganisms that have adapted to thrive on plastic as they may contain genes, enzymes or pathways involved in the adhesion or metabolism of plastics. We implemented DNA metabarcoding with nanopore MinION sequencing to compare the 1-month-old biomes of hydrolyzable (polyethylene terephthalate) and non-hydrolyzable (polyethylene) plastics surfaces vs. ...[more]

PMID: 35495680

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Project description:BackgroundThe biological degradation of plastics is a promising method to counter the increasing pollution of our planet with artificial polymers and to develop eco-friendly recycling strategies. Polyethylene terephthalate (PET) is a thermoplast industrially produced from fossil feedstocks since the 1940s, nowadays prevalently used in bottle packaging and textiles. Although established industrial processes for PET recycling exist, large amounts of PET still end up in the environment-a significant portion thereof in the world's oceans. In 2016, Ideonella sakaiensis, a bacterium possessing the ability to degrade PET and use the degradation products as a sole carbon source for growth, was isolated. I. sakaiensis expresses a key enzyme responsible for the breakdown of PET into monomers: PETase. This hydrolase might possess huge potential for the development of biological PET degradation and recycling processes as well as bioremediation approaches of environmental plastic waste.ResultsUsing the photosynthetic microalga Phaeodactylum tricornutum as a chassis we generated a microbial cell factory capable of producing and secreting an engineered version of PETase into the surrounding culture medium. Initial degradation experiments using culture supernatant at 30 °C showed that PETase possessed activity against PET and the copolymer polyethylene terephthalate glycol (PETG) with an approximately 80-fold higher turnover of low crystallinity PETG compared to bottle PET. Moreover, we show that diatom produced PETase was active against industrially shredded PET in a saltwater-based environment even at mesophilic temperatures (21 °C). The products resulting from the degradation of the PET substrate were mainly terephthalic acid (TPA) and mono(2-hydroxyethyl) terephthalic acid (MHET) estimated to be formed in the micromolar range under the selected reaction conditions.ConclusionWe provide a promising and eco-friendly solution for biological decomposition of PET waste in a saltwater-based environment by using a eukaryotic microalga instead of a bacterium as a model system. Our results show that via synthetic biology the diatom P. tricornutum indeed could be converted into a valuable chassis for biological PET degradation. Overall, this proof of principle study demonstrates the potential of the diatom system for future biotechnological applications in biological PET degradation especially for bioremediation approaches of PET polluted seawater.

Project description:Biodegradation of terephthalate (TPA) is a highly desired catabolic process for the bacterial utilization of this polyethylene terephthalate (PET) depolymerization product, but to date, the structure of terephthalate dioxygenase (TPDO), a Rieske oxygenase (RO) that catalyzes the dihydroxylation of TPA to a cis-diol, is unavailable. In this study, we characterized the steady-state kinetics and first crystal structure of TPDO from Comamonas testosteroni KF1 (TPDOKF1). TPDOKF1 exhibited substrate specificity for TPA (kcat/Km = 57 ± 9 mM-1 s-1). The TPDOKF1 structure harbors characteristic RO features as well as a unique catalytic domain that rationalizes the enzyme's function. The docking and mutagenesis studies reveal that its substrate specificity for TPA is mediated by the Arg309 and Arg390 residues, positioned on opposite faces of the active site. Additionally, residue Gln300 is also proven to be crucial for the activity, as its mutation to alanine decreases the activity (kcat) by 80%. This study delineates the structural features that dictate the substrate recognition and specificity of TPDO. IMPORTANCE Global plastic pollution has become the most pressing environmental issue. Recent studies on enzymes depolymerizing polyethylene terephthalate plastic into terephthalate (TPA) show some potential for tackling this. Microbial utilization of this released product, TPA, is an emerging and promising strategy for waste-to-value creation. Research in the last decade has identified terephthalate dioxygenase (TPDO) as being responsible for initiating the enzymatic degradation of TPA in a few Gram-negative and Gram-positive bacteria. Here, we determined the crystal structure of TPDO from Comamonas testosteroni KF1 and revealed that it possesses a unique catalytic domain featuring two basic residues in the active site to recognize TPA. Biochemical and mutagenesis studies demonstrated the crucial residues responsible for the substrate specificity of this enzyme.

Dataset Information

High-Resolution Screening for Marine Prokaryotes and Eukaryotes With Selective Preference for Polyethylene and Polyethylene Terephthalate Surfaces.

Publications

High-Resolution Screening for Marine Prokaryotes and Eukaryotes With Selective Preference for Polyethylene and Polyethylene Terephthalate Surfaces.

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