Characterization and engineering of a two-enzyme system for plastics depolymerization.
13 Oct 2020-Proceedings of the National Academy of Sciences of the United States of America (National Academy of Sciences)-Vol. 117, Iss: 41, pp 25476-25485
TL;DR: The characterization of the MHETase enzyme and synergy of the two-enzyme PET depolymerization system may inform enzyme cocktail-based strategies for plastics upcycling and will inform future efforts in the biological deconstruction andUpcycling of mixed plastics.
Abstract: Plastics pollution represents a global environmental crisis. In response, microbes are evolving the capacity to utilize synthetic polymers as carbon and energy sources. Recently, Ideonella sakaiensis was reported to secrete a two-enzyme system to deconstruct polyethylene terephthalate (PET) to its constituent monomers. Specifically, the I. sakaiensis PETase depolymerizes PET, liberating soluble products, including mono(2-hydroxyethyl) terephthalate (MHET), which is cleaved to terephthalic acid and ethylene glycol by MHETase. Here, we report a 1.6 A resolution MHETase structure, illustrating that the MHETase core domain is similar to PETase, capped by a lid domain. Simulations of the catalytic itinerary predict that MHETase follows the canonical two-step serine hydrolase mechanism. Bioinformatics analysis suggests that MHETase evolved from ferulic acid esterases, and two homologous enzymes are shown to exhibit MHET turnover. Analysis of the two homologous enzymes and the MHETase S131G mutant demonstrates the importance of this residue for accommodation of MHET in the active site. We also demonstrate that the MHETase lid is crucial for hydrolysis of MHET and, furthermore, that MHETase does not turnover mono(2-hydroxyethyl)-furanoate or mono(2-hydroxyethyl)-isophthalate. A highly synergistic relationship between PETase and MHETase was observed for the conversion of amorphous PET film to monomers across all nonzero MHETase concentrations tested. Finally, we compare the performance of MHETase:PETase chimeric proteins of varying linker lengths, which all exhibit improved PET and MHET turnover relative to the free enzymes. Together, these results offer insights into the two-enzyme PET depolymerization system and will inform future efforts in the biological deconstruction and upcycling of mixed plastics.
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TL;DR: The AlphaFold Protein Structure Database (AlphaFold DB, https://alphafold.ebi.ac.uk) is an openly accessible, extensive database of high-accuracy protein-structure predictions.
Abstract: The AlphaFold Protein Structure Database (AlphaFold DB, https://alphafold.ebi.ac.uk) is an openly accessible, extensive database of high-accuracy protein-structure predictions. Powered by AlphaFold v2.0 of DeepMind, it has enabled an unprecedented expansion of the structural coverage of the known protein-sequence space. AlphaFold DB provides programmatic access to and interactive visualization of predicted atomic coordinates, per-residue and pairwise model-confidence estimates and predicted aligned errors. The initial release of AlphaFold DB contains over 360,000 predicted structures across 21 model-organism proteomes, which will soon be expanded to cover most of the (over 100 million) representative sequences from the UniRef90 data set.
2,008 citations
01 Jul 2021
TL;DR: In this article, the challenges and opportunities associated with the catalytic transformation of waste plastics, looking at both chemical and biological approaches to transforming such spent materials into a resource, are explored and compared.
Abstract: Plastics pollution is causing an environmental crisis, prompting the development of new approaches for recycling, and upcycling. Here, we review challenges and opportunities in chemical and biological catalysis for plastics deconstruction, recycling, and upcycling. We stress the need for rigorous characterization and use of widely available substrates, such that catalyst performance can be compared across studies. Where appropriate, we draw parallels between catalysis on biomass and plastics, as both substrates are low-value, solid, recalcitrant polymers. Innovations in catalyst design and reaction engineering are needed to overcome kinetic and thermodynamic limitations of plastics deconstruction. Either chemical and biological catalysts will need to act interfacially, where catalysts function at a solid surface, or polymers will need to be solubilized or processed to smaller intermediates to facilitate improved catalyst–substrate interaction. Overall, developing catalyst-driven technologies for plastics deconstruction and upcycling is critical to incentivize improved plastics reclamation and reduce the severe global burden of plastic waste. Plastics are invaluable materials for modern society, although they result in the generation of large amounts of litter at the end of their life cycle. This Review explores the challenges and opportunities associated with the catalytic transformation of waste plastics, looking at both chemical and biological approaches to transforming such spent materials into a resource.
243 citations
TL;DR: In this article, an improved deterministic kinetic model for the dominating reaction families of solid plastic waste (SPW) was proposed to identify the leading recycling technologies, minimizing the global warming potential in an industrial context.
214 citations
TL;DR: In this paper, the authors present process modeling, techno-economic, life-cycle, and socioeconomic impact analyses for an enzymatic PET depolymerization-based recycling process, which they compare with virgin TPA manufacturing.
100 citations
TL;DR: In this paper, the degradation of microplastics is closely related to the enzymatic reactions produced by the microorganisms, such as algae, fungi, and bacteria, and the degradation mechanisms remain unclear.
Abstract: Microplastics contamination is becoming a major concern worldwide. More than 1 million seabirds and 100,000 sea animals have died due to plastic contamination. In addition, plastic particles have been found in juvenile turtles. Statistical data on plastic pollution indicate that this is a serious issue. Due to their small size, microplastics have a large surface area and have more ability to absorb into biological cells. The hydrophobic surface of microplastics attracts co-contaminants such as heavy metals, pharmaceutical toxicants, flame retardants, and other plasticizers, which can then enter biological organisms. Microplastics are usually recalcitrant in the environment, causing microplastics to be transported along the food chain, with humans as the final consumer. Research has been conducted to evaluate the best way to treat and remediate microplastic pollution. Research on microplastic degradation is focused on biological and non-biological approaches. To date, microorganisms such as algae, fungi, and bacteria have attracted the attention of scientists as a tool for microplastic treatment. The degradation of microplastics is closely related to the enzymatic reactions produced by the microorganisms. Here we review microplastics degradation through enzymes from the microorganism’s perspective. We present the enzymes that have been isolated from microorganisms for specific microplastics; the mechanisms of microplastics degradation by various enzymes; and the types of microplastics for which degradation mechanisms remain unclear.
93 citations
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Abstract: The BLAST programs are widely used tools for searching protein and DNA databases for sequence similarities. For protein comparisons, a variety of definitional, algorithmic and statistical refinements described here permits the execution time of the BLAST programs to be decreased substantially while enhancing their sensitivity to weak similarities. A new criterion for triggering the extension of word hits, combined with a new heuristic for generating gapped alignments, yields a gapped BLAST program that runs at approximately three times the speed of the original. In addition, a method is introduced for automatically combining statistically significant alignments produced by BLAST into a position-specific score matrix, and searching the database using this matrix. The resulting Position-Specific Iterated BLAST (PSIBLAST) program runs at approximately the same speed per iteration as gapped BLAST, but in many cases is much more sensitive to weak but biologically relevant sequence similarities. PSI-BLAST is used to uncover several new and interesting members of the BRCT superfamily.
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TL;DR: NAMD as discussed by the authors is a parallel molecular dynamics code designed for high-performance simulation of large biomolecular systems that scales to hundreds of processors on high-end parallel platforms, as well as tens of processors in low-cost commodity clusters, and also runs on individual desktop and laptop computers.
Abstract: NAMD is a parallel molecular dynamics code designed for high-performance simulation of large biomolecular systems. NAMD scales to hundreds of processors on high-end parallel platforms, as well as tens of processors on low-cost commodity clusters, and also runs on individual desktop and laptop computers. NAMD works with AMBER and CHARMM potential functions, parameters, and file formats. This article, directed to novices as well as experts, first introduces concepts and methods used in the NAMD program, describing the classical molecular dynamics force field, equations of motion, and integration methods along with the efficient electrostatics evaluation algorithms employed and temperature and pressure controls used. Features for steering the simulation across barriers and for calculating both alchemical and conformational free energy differences are presented. The motivations for and a roadmap to the internal design of NAMD, implemented in C++ and based on Charm++ parallel objects, are outlined. The factors affecting the serial and parallel performance of a simulation are discussed. Finally, typical NAMD use is illustrated with representative applications to a small, a medium, and a large biomolecular system, highlighting particular features of NAMD, for example, the Tcl scripting language. The article also provides a list of the key features of NAMD and discusses the benefits of combining NAMD with the molecular graphics/sequence analysis software VMD and the grid computing/collaboratory software BioCoRE. NAMD is distributed free of charge with source code at www.ks.uiuc.edu.
14,558 citations
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...To examine MHETase dynamics and ligand stability, classic molecular dynamics (MD) simulations were conducted with NAMD (40) (all simulations totaling 2.25 μs) utilizing the CHARMM forcefield (41)....
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7,672 citations
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...We utilized the Amber software package (42) to perform hybrid quantum mechanics/molecular mechanics (QM/MM) 2D umbrella sampling with semiempirical force field SCC-DFTB (43) to study the catalytic steps....
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...Classic MD simulations were run with NAMD (40); QM/MM simulations, including 2D umbrella sampling free-energy calculations, were run in Amber (42, 72)....
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