8qrj: Difference between revisions
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==LCC-ICCG PETase mutant H218Y== | |||
<StructureSection load='8qrj' size='340' side='right'caption='[[8qrj]], [[Resolution|resolution]] 1.42Å' scene=''> | |||
== Structural highlights == | |||
<table><tr><td colspan='2'>[[8qrj]] is a 1 chain structure with sequence from [https://en.wikipedia.org/wiki/Unidentified_prokaryotic_organism Unidentified prokaryotic organism]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=8QRJ OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=8QRJ FirstGlance]. <br> | |||
</td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 1.42Å</td></tr> | |||
<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CA:CALCIUM+ION'>CA</scene>, <scene name='pdbligand=EDO:1,2-ETHANEDIOL'>EDO</scene>, <scene name='pdbligand=P33:3,6,9,12,15,18-HEXAOXAICOSANE-1,20-DIOL'>P33</scene></td></tr> | |||
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=8qrj FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=8qrj OCA], [https://pdbe.org/8qrj PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=8qrj RCSB], [https://www.ebi.ac.uk/pdbsum/8qrj PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=8qrj ProSAT]</span></td></tr> | |||
</table> | |||
== Function == | |||
[https://www.uniprot.org/uniprot/PETH_UNKP PETH_UNKP] Catalyzes the hydrolysis of cutin, a polyester that forms the structure of plant cuticle (PubMed:22194294). Shows esterase activity towards p-nitrophenol-linked aliphatic esters (pNP-aliphatic esters), with a preference for short-chain substrates (C4 substrate at most) (PubMed:22194294, PubMed:24593046). Cannot hydrolyze olive oil (PubMed:22194294). Is also able to degrade poly(ethylene terephthalate), the most abundant polyester plastic in the world (PubMed:22194294, PubMed:32269349). Can also depolymerize poly(epsilon-caprolactone) (PCL), a synthetic aliphatic biodegradable polyester (PubMed:22194294).<ref>PMID:22194294</ref> <ref>PMID:24593046</ref> <ref>PMID:32269349</ref> | |||
<div style="background-color:#fffaf0;"> | |||
== Publication Abstract from PubMed == | |||
Enzyme-mediated polyethylene terephthalate (PET) depolymerization has recently emerged as a sustainable solution for PET recycling. Towards an industrial-scale implementation of this technology, various strategies are being explored to enhance PET depolymerization (PETase) activity and improve enzyme stability, expression, and purification processes. Recently, rational engineering of a known PET hydrolase (LCC-leaf compost cutinase) has resulted in the isolation of a variant harboring four-point mutations (LCC-ICCG), presenting increased PETase activity and thermal stability. Here, we revealed the enzyme's natural extracellular expression and used it to efficiently screen error-prone genetic libraries based on LCC-ICCG for enhanced activity toward consumer-grade PET. Following multiple rounds of mutagenesis and screening, we successfully isolated variants that exhibited up to a 60% increase in PETase activity. Among other mutations, the improved variants showed a histidine to tyrosine substitution at position 218, a residue known to be involved in substrate binding and stabilization. Introducing H218Y mutation on the background of LCC-ICCG (named here LCC-ICCG/H218Y) resulted in a similar level of activity improvement. Analysis of the solved structure of LCC-ICCG/H218Y compared to other known PETases featuring different amino acids at the equivalent position suggests that H218Y substitution promotes enhanced PETase activity. The expression and screening processes developed in this study can be further used to optimize additional enzymatic parameters crucial for efficient enzymatic degradation of consumer-grade PET. | |||
Streamlined screening of extracellularly expressed PETase libraries for improved polyethylene terephthalate degradation.,Orr G, Niv Y, Barakat M, Boginya A, Dessau M, Afriat-Jurnou L Biotechnol J. 2024 Jul;19(7):e2400021. doi: 10.1002/biot.202400021. PMID:38987219<ref>PMID:38987219</ref> | |||
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | |||
[[Category: | </div> | ||
[[Category: | <div class="pdbe-citations 8qrj" style="background-color:#fffaf0;"></div> | ||
[[Category: Barakat | == References == | ||
[[Category: | <references/> | ||
[[Category: | __TOC__ | ||
[[Category: Niv | </StructureSection> | ||
[[Category: | [[Category: Large Structures]] | ||
[[Category: Unidentified prokaryotic organism]] | |||
[[Category: Afriat-Jurnou L]] | |||
[[Category: Barakat M]] | |||
[[Category: Boginya A]] | |||
[[Category: Dessau M]] | |||
[[Category: Niv Y]] | |||
[[Category: Orr G]] | |||
Latest revision as of 07:51, 18 September 2024
LCC-ICCG PETase mutant H218YLCC-ICCG PETase mutant H218Y
Structural highlights
FunctionPETH_UNKP Catalyzes the hydrolysis of cutin, a polyester that forms the structure of plant cuticle (PubMed:22194294). Shows esterase activity towards p-nitrophenol-linked aliphatic esters (pNP-aliphatic esters), with a preference for short-chain substrates (C4 substrate at most) (PubMed:22194294, PubMed:24593046). Cannot hydrolyze olive oil (PubMed:22194294). Is also able to degrade poly(ethylene terephthalate), the most abundant polyester plastic in the world (PubMed:22194294, PubMed:32269349). Can also depolymerize poly(epsilon-caprolactone) (PCL), a synthetic aliphatic biodegradable polyester (PubMed:22194294).[1] [2] [3] Publication Abstract from PubMedEnzyme-mediated polyethylene terephthalate (PET) depolymerization has recently emerged as a sustainable solution for PET recycling. Towards an industrial-scale implementation of this technology, various strategies are being explored to enhance PET depolymerization (PETase) activity and improve enzyme stability, expression, and purification processes. Recently, rational engineering of a known PET hydrolase (LCC-leaf compost cutinase) has resulted in the isolation of a variant harboring four-point mutations (LCC-ICCG), presenting increased PETase activity and thermal stability. Here, we revealed the enzyme's natural extracellular expression and used it to efficiently screen error-prone genetic libraries based on LCC-ICCG for enhanced activity toward consumer-grade PET. Following multiple rounds of mutagenesis and screening, we successfully isolated variants that exhibited up to a 60% increase in PETase activity. Among other mutations, the improved variants showed a histidine to tyrosine substitution at position 218, a residue known to be involved in substrate binding and stabilization. Introducing H218Y mutation on the background of LCC-ICCG (named here LCC-ICCG/H218Y) resulted in a similar level of activity improvement. Analysis of the solved structure of LCC-ICCG/H218Y compared to other known PETases featuring different amino acids at the equivalent position suggests that H218Y substitution promotes enhanced PETase activity. The expression and screening processes developed in this study can be further used to optimize additional enzymatic parameters crucial for efficient enzymatic degradation of consumer-grade PET. Streamlined screening of extracellularly expressed PETase libraries for improved polyethylene terephthalate degradation.,Orr G, Niv Y, Barakat M, Boginya A, Dessau M, Afriat-Jurnou L Biotechnol J. 2024 Jul;19(7):e2400021. doi: 10.1002/biot.202400021. PMID:38987219[4] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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