PETase

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PETase
PETase 5XH3 with HEMT-cartoon.png
I. sakaiensis PETase ( A0A0K8P6T7 ) in complex with HEMT, a PET analogue ( PDB: 5XH3 ​).
Identifiers
EC no. 3.1.1.101
Alt. namesPET hydrolase, poly(ethylene terephthalate) hydrolase
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PETases are an esterase class of enzymes that catalyze the breakdown (via hydrolysis) of polyethylene terephthalate (PET) plastic to monomeric mono-2-hydroxyethyl terephthalate (MHET). The idealized chemical reaction is:

Contents

(ethylene terephthalate)n + H2O → (ethylene terephthalate)n-1 + MHET,

where n is the number of monomers in the polymer chain, though a trace amount of the PET breaks down instead to bis(2-hydroxyethyl) terephthalate (BHET). [1] PETases can also break down PEF-plastic (polyethylene-2,5-furandicarboxylate), which is a bioderived PET replacement, into the analogous MHEF. PETases can't catalyze the hydrolysis of aliphatic polyesters like polybutylene succinate or polylactic acid. [2]

Whereas the degradation of PET by natural (non-enzymatic) means will take hundreds of years, PETases can degrade it in a matter of days. [3]

History

The first PETase was discovered in 2016 from Ideonella sakaiensis strain 201-F6 bacteria found from sludge samples collected close to a Japanese PET bottle recycling site. [1] [4] There were other types of hydrolases previously known to degrade PET, [2] including lipases, esterases, and cutinases. [5] For comparison, enzymes that degrade polyester have been known to exist at least as far back as 1975 (in the case of α-chymotrypsin) [6] and 1977 (lipase). [7]

PET plastic came into widespread use in the 1970s and it has been suggested that PETases in bacteria evolved only recently. [2] PETase may have had past enzymatic activity associated with degradation of a waxy coating on plants. [8]

Structure

As of April 2019, there were 17 known three-dimensional crystal structures of PETases: 6QGC, 6ILX, 6ILW, 5YFE, 6EQD, 6EQE, 6EQF, 6EQG, 6EQH, 6ANE, 5XJH, 5YNS, 5XFY, 5XFZ, 5XG0, 5XH2 and 5XH3.

PETase exhibits shared qualities with both lipases and cutinases in that it possesses an α/β-hydrolase fold; although, the active-site cleft observed in PETase is more open than in cutinases. [2] The Ideonella sakaiensis PETase is similar to dienelactone hydrolase, according to Pfam. According to ESTHER, it falls into the Polyesterase-lipase-cutinase family.

There are approximately 69 PETase-like enzymes comprising a variety of diverse organisms, and there are two classifications of these enzymes including type I and type II. It is suggested that 57 enzymes fall into the type I category whereas the rest fall into the type II group, including the PETase enzyme found in the Ideonella sakaiensis. Within all 69 PETase-like enzymes, there exists the same three residues within the active site, suggesting that the catalytic mechanism is the same in all forms of PETase-like enzymes. [9]

Mutations

The discovery of PETase from I. sakaiensis provides a potential solution to the world’s amassing plastic; however, naturally occurring enzymes are limited in their degradation abilities due to instability, low activity, and expression levels, which ultimately drive the need for improvement if they are to be used for large-scale industrial applications. [10] The majority of strategies implement site-directed mutagenesis to create an improved version, known as a variant or mutant, of the enzyme. One variant increased the activity of PETase by 22.4% by replacing the arginine with alanine in the amino acid chain at the 280th position. [11] Similarly, a double mutant was created to constrict the active site and became 4.13% more active than the wildtype. [12] Comparatively, two other double mutants created extra hydrogen bonds that improved the stability of PETase. Other successful approaches to improving PETase stability include adding Ca2+ or Mg2+, disulfide bonds and salt bridges as well as glycosylation. [10] For thermal stability, another double mutant displayed an increase in comparison to the wild type. [11] Moreover, the β1-β2 connecting loop of the enzyme may also be a future target for improved thermal stability due to its flexibility and distance from the active site. [13]

Biological pathway

PETase and MHETase reaction pathway. Plastic Breakdown by PETase.png
PETase and MHETase reaction pathway.

In I. sakaiensis, the resultant MHET is further broken down by the action of MHETase enzyme to terephthalic acid and ethylene glycol. [1] Laboratory experiments showed that chimeric proteins that artificially link a MHETase and a PETase outperform similar mixtures of free enzymes. [15]

See also

Related Research Articles

Transesterification is the process of exchanging the organic functional group R″ of an ester with the organic group R' of an alcohol. These reactions are often catalyzed by the addition of an acid or base catalyst. Strong acids catalyze the reaction by donating a proton to the carbonyl group, thus making it a more potent electrophile. Bases catalyze the reaction by removing a proton from the alcohol, thus making it more nucleophilic. The reaction can also be accomplished with the help of other enzymes, particularly lipases.

<span class="mw-page-title-main">Polyethylene terephthalate</span> Polymer

Polyethylene terephthalate (or poly(ethylene terephthalate), PET, PETE, or the obsolete PETP or PET-P), is the most common thermoplastic polymer resin of the polyester family and is used in fibres for clothing, containers for liquids and foods, and thermoforming for manufacturing, and in combination with glass fibre for engineering resins.

In biochemistry, hydrolases constitute a class of enzymes that commonly function as biochemical catalysts that use water to break a chemical bond:

<span class="mw-page-title-main">PET bottle recycling</span> Recycling of bottles made of polyethylene terephthalate

Although PET is used in several applications,, as of 2022 only bottles are collected at a substantial scale. The main motivations have been either cost reduction or recycle content of retail goods. An increasing amount is recycled back into bottles, the rest goes into fibres, film, thermoformed packaging and strapping. After sorting, cleaning and grinding, 'bottle flake' is obtained, which is then processed by either:

<span class="mw-page-title-main">Polyethylene naphthalate</span> Chemical compound

Polyethylene naphthalate is a polyester derived from naphthalene-2,6-dicarboxylic acid and ethylene glycol. As such it is related to poly(ethylene terephthalate), but with superior barrier properties.

<span class="mw-page-title-main">Polyester</span> Category of polymers, in which the monomers are joined together by ester links

Polyester is a category of polymers that contain the ester functional group in every repeat unit of their main chain. As a specific material, it most commonly refers to a type called polyethylene terephthalate (PET). Polyesters include naturally occurring chemicals, such as in plants and insects, as well as synthetics such as polybutyrate. Natural polyesters and a few synthetic ones are biodegradable, but most synthetic polyesters are not. Synthetic polyesters are used extensively in clothing.

<span class="mw-page-title-main">Monoacylglycerol lipase</span> Class of enzymes

Monoacylglycerol lipase is an enzyme that, in humans, is encoded by the MGLL gene. MAGL is a 33-kDa, membrane-associated member of the serine hydrolase superfamily and contains the classical GXSXG consensus sequence common to most serine hydrolases. The catalytic triad has been identified as Ser122, His269, and Asp239.

<span class="mw-page-title-main">Chlorophyllase</span> Enzyme in chlorophyll metabolism

Chlorophyllase is an essential enzyme in chlorophyll metabolism. It is a membrane proteins commonly known as chlase (EC 3.1.1.14, CLH) with systematic name chlorophyll chlorophyllidohydrolase. It catalyzes the reaction

<span class="mw-page-title-main">Cutinase</span> Class of enzymes

The enzyme cutinase is a member of the hydrolase family. It catalyzes the following reaction:

The enzyme poly(3-hydroxyoctanoate) depolymerase (EC 3.1.1.76) catalyzes the hydrolysis of the polyester poly{oxycarbonyl[(R)-2-pentylethylene] to oligomers

Biodegradable additives are additives that enhance the biodegradation of polymers by allowing microorganisms to utilize the carbon within the polymer chain as a source of energy. Biodegradable additives attract microorganisms to the polymer through quorum sensing after biofilm creation on the plastic product. Additives are generally in masterbatch formation that use carrier resins such as polyethylene (PE), polypropylene (PP), polystyrene (PS) or polyethylene terephthalate (PET).

Ideonella is a genus of bacteria in the family Comamonadaceae.

Poly(ethylene adipate) Chemical compound

Poly(ethylene adipate) or PEA is an aliphatic polyester. It is most commonly synthesized from a polycondensation reaction between ethylene glycol and adipic acid. PEA has been studied as it is biodegradable through a variety of mechanisms and also fairly inexpensive compared to other polymers. Its lower molecular weight compared to many polymers aids in its biodegradability.

<span class="mw-page-title-main">Edible packaging</span>

Edible packaging refers to packaging which is edible and biodegradable.

Ideonella sakaiensis is a bacterium from the genus Ideonella and family Comamonadaceae capable of breaking down and consuming the plastic polyethylene terephthalate (PET) using it as both a carbon and energy source. The bacterium was originally isolated from a sediment sample taken outside of a plastic bottle recycling facility in Sakai City, Japan.

<span class="mw-page-title-main">John McGeehan</span> British research scientist

John McGeehan is a British research scientist and professor of structural biology. He was director of the Centre for Enzyme Innovation (CEI) at the University of Portsmouth until 2022 and led a research team on enzyme engineering.

2-Hydroxyethyl terephthalic acid is an organic compound with the formula HOC2H4O2CC6H4CO2H. It is the monoester of terephthalic acid and ethylene glycol. The compound is a precursor to poly(ethylene terephthalate) (PET), a polymer that is produced on a large scale industrially. 2-Hydroxyethyl terephthalic acid is a colorless solid that is soluble in water and polar organic solvents. Near neutral pH, 2-hydroxyethyl terephthalic acid converts to 2-hydroxyethyl terephthalate, HOC2H4O2CC6H4CO2−.

Kohei Oda is a Japanese microbiologist and an emeritus professor at Kyoto Institute of Technology. He is known for his work on bacterial discovery and bacterial metabolism. In particular, he led a team of Japanese scientists in the discovery of plastic-degrading bacteria, Ideonella sakaiensis, in 2016.

<span class="mw-page-title-main">MHETase</span>

The enzyme MHETase is a hydrolase, which was discovered in 2016. It cleaves 2-hydroxyethyl terephthalic acid, the PET degradation product by PETase, to ethylene glycol and terephthalic acid. This pair of enzymes, PETase and MHETase, enable the bacterium Ideonella sakaiensis to live on the plastic PET as sole carbon source.

<span class="mw-page-title-main">Plastic degradation by marine bacteria</span> Ability of bacteria to break down plastic polymers

Plastic degradation in marine bacteria describes when certain pelagic bacteria break down polymers and use them as a primary source of carbon for energy. Polymers such as polyethylene(PE), polypropylene (PP), and polyethylene terephthalate (PET) are incredibly useful for their durability and relatively low cost of production, however it is their persistence and difficulty to be properly disposed of that is leading to pollution of the environment and disruption of natural processes. It is estimated that each year there are 9-14 million metric tons of plastic that are entering the ocean due to inefficient solutions for their disposal. The biochemical pathways that allow for certain microbes to break down these polymers into less harmful byproducts has been a topic of study to develop a suitable anti-pollutant.

References

  1. 1 2 3 Yoshida S, Hiraga K, Takehana T, Taniguchi I, Yamaji H, Maeda Y, et al. (March 2016). "A bacterium that degrades and assimilates poly(ethylene terephthalate)". Science. 351 (6278): 1196–9. doi:10.1126/science.aad6359. PMID   26965627. S2CID   31146235.
  2. 1 2 3 4 Austin HP, Allen MD, Donohoe BS, Rorrer NA, Kearns FL, Silveira RL, et al. (May 2018). "Characterization and engineering of a plastic-degrading aromatic polyesterase". Proceedings of the National Academy of Sciences of the United States of America. 115 (19): E4350–E4357. doi: 10.1073/pnas.1718804115 . PMC   5948967 . PMID   29666242.
  3. ↑ Dockrill, Peter. "Scientists Have Accidentally Created a Mutant Enzyme That Eats Plastic Waste". ScienceAlert. Retrieved 2018-11-27.
  4. ↑ Tanasupawat S, Takehana T, Yoshida S, Hiraga K, Oda K (August 2016). "Ideonella sakaiensis sp. nov., isolated from a microbial consortium that degrades poly(ethylene terephthalate)". International Journal of Systematic and Evolutionary Microbiology. 66 (8): 2813–8. doi: 10.1099/ijsem.0.001058 . PMID   27045688.
  5. ↑ Han X, Liu W, Huang JW, Ma J, Zheng Y, Ko TP, et al. (December 2017). "Structural insight into catalytic mechanism of PET hydrolase". Nature Communications. 8 (1): 2106. doi:10.1038/s41467-017-02255-z. PMC   5727383 . PMID   29235460.
  6. ↑ Tabushi I, Yamada H, Matsuzaki H, Furukawa J (August 1975). "Polyester readily hydrolyzable by chymotrypsin". Journal of Polymer Science: Polymer Letters Edition. 13 (8): 447–450. doi:10.1002/pol.1975.130130801.
  7. ↑ Tokiwa Y, Suzuki T (November 1977). "Hydrolysis of polyesters by lipases". Nature. 270 (5632): 76–8. doi:10.1038/270076a0. PMID   927523. S2CID   4145159.
  8. ↑ "Lab 'Accident' Becomes Mutant Enzyme That Devours Plastic". Live Science. Retrieved 2018-11-27.
  9. 1 2 Joo S, Cho IJ, Seo H, Son HF, Sagong HY, Shin TJ, et al. (January 2018). "Structural insight into molecular mechanism of poly(ethylene terephthalate) degradation". Nature Communications. 9 (1): 382. doi:10.1038/s41467-018-02881-1. PMC   5785972 . PMID   29374183.
  10. 1 2 Qi, Xinhua; Yan, Wenlong; Cao, Zhibei; Ding, Mingzhu; Yuan, Yingjin (2021-12-26). "Current Advances in the Biodegradation and Bioconversion of Polyethylene Terephthalate". Microorganisms. 10 (1): 39. doi:10.3390/microorganisms10010039. ISSN   2076-2607. PMC   8779501 . PMID   35056486.{{cite journal}}: CS1 maint: unflagged free DOI (link)
  11. 1 2 Urbanek, Aneta K.; Kosiorowska, Katarzyna E.; Mirończuk, Aleksandra M. (2021-11-30). "Current Knowledge on Polyethylene Terephthalate Degradation by Genetically Modified Microorganisms". Frontiers in Bioengineering and Biotechnology. 9. doi:10.3389/fbioe.2021.771133. ISSN   2296-4185. PMC   8669999 . PMID   34917598.{{cite journal}}: CS1 maint: unflagged free DOI (link)
  12. ↑ Austin, Harry P.; Allen, Mark D.; Donohoe, Bryon S.; Rorrer, Nicholas A.; Kearns, Fiona L.; Silveira, Rodrigo L.; Pollard, Benjamin C.; Dominick, Graham; Duman, Ramona; El Omari, Kamel; Mykhaylyk, Vitaliy; Wagner, Armin; Michener, William E.; Amore, Antonella; Skaf, Munir S. (2018-05-08). "Characterization and engineering of a plastic-degrading aromatic polyesterase". Proceedings of the National Academy of Sciences. 115 (19). doi:10.1073/pnas.1718804115. ISSN   0027-8424. PMC   5948967 . PMID   29666242.
  13. ↑ da Costa, Clauber Henrique Souza; dos Santos, Alberto M.; Alves, Cláudio Nahum; Martí, Sérgio; Moliner, Vicent; Santana, Kauê; Lameira, Jerônimo (October 2021). "Assessment of the PETase conformational changes induced by poly(ethylene terephthalate) binding". Proteins: Structure, Function, and Bioinformatics. 89 (10): 1340–1352. doi:10.1002/prot.26155. ISSN   0887-3585.
  14. ↑ Allison Chan (2016). "The Future of Bacteria Cleaning Our Plastic Waste" (PDF).
  15. ↑ Knott BC, Erickson E, Allen MD, Gado JE, Graham R, Kearns FL, et al. (Oct 2020). "Characterization and engineering of a two-enzyme system for plastics depolymerization". Proc Natl Acad Sci U S A. 117 (41): 25476–25485. doi: 10.1073/pnas.2006753117 . PMC   7568301 . PMID   32989159.
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