Files in this item
Computational insights into catalytic mechanism and thermostability of the enzyme Is-PETase
|Mitchell, John B. O.
|The field of study surrounding the PETase class of enzymes has gained a great deal of popularity in the past five years. PETases are enzymes capable of degrading poly(ethylene) terephthalate and presents an opportunity to bioanalytically recycle this common pollutant. The most extensively researched PETase is the enzyme Is-PETase, which originates from the organism Idonella sakaiensis and was discovered near a recycling plant in 2016. To further the utility of Is-PETase in the industrial recycling of poly(ethylene) terephthalate, improvements in desirable properties such as catalytic rate and thermostability must be achieved via introduction of amino-acid substitutions to the enzyme. In order to further improve the activity of Is-PETase, it would be informative to have a deeper understanding of its catalytic mechanism. To this end, we have investigated the catalytic mechanism of the degradation of poly(ethylene) terephthalate via Is-PETase. We applied hybrid quantum mechanical/ molecular mechanical to generate several independent reaction profiles. From these reaction profiles we have concluded that this reaction proceeds with an overall activation barrier of 35.6 kJ mol-1 . This relatively low activation barrier suggests that the ‘true’ rate-limiting step for this reaction is a physical process such as substrate binding or product dissociation. Our calculations provide preliminary evidence for the product dissociation step being rate-limiting. We have also investigated modifications to wild-type Is-PETase that contribute to the enzyme’s thermostability. We have created our own mutant of Is-PETase, DISU-PETase, through the in silico introduction of a novel disulfide bond. We generated reaction profiles for DISU-PETase using the same quantum mechanical/ molecular mechanical techniques as applied to wild-type Is-PETase. We found that introduction of the novel disulfide bond in DISU-PETase had no adverse effect upon the reaction profile of the degradation of poly(ethylene) terephthalate. We have also applied molecular dynamics techniques to investigate the intramolecular interactions that contribute toward the thermostability of the most highly active variants of Is-PETase in the literature. We have also used our quantum mechanical/ molecular mechanical techniques to generate reaction profiles for the degradation of an alternate substrate, poly(ethylene) furanoate via wildtype Is-PETase.
|Computational Insights into the Catalytic Mechanism of Is-PETase: An Enzyme Capable of degrading poly(ethylene) terephthalate (thesis data) Shrimpton-Phoenix, E., Mitchell, J. B. O., and Buehl, M., University of St Andrews, 27 Oct 2022. DOI: 10.17630/486acc90-3be5-43c2-aa08-17c02096bb85
|Computational insights into catalytic mechanism and thermostability of the enzyme Is-PETase
|PhD Doctor of Philosophy
|The University of St Andrews
This item appears in the following Collection(s)
Items in the St Andrews Research Repository are protected by copyright, with all rights reserved, unless otherwise indicated.