Journal
CHEMPHYSCHEM
Volume 23, Issue 20, Pages -Publisher
WILEY-V C H VERLAG GMBH
DOI: 10.1002/cphc.202200269
Keywords
glucuronoyl esterase; lignocellulose processing; enzymatic catalysis; QM/MM; thermophilic enzyme
Funding
- FCT/MCTES [UIBD/50006/2020, PTDC/QUI-QFI/28714/2017, PTDC/QUIOUT/1401/2020]
- FCT/MCTES (Fundacao para a Ciencia e Tecnologia/Ministerio da Ciencia, Tecnologia e Ensino Superior)
- ESF (European Social Fund) through POCH (Programa Operacional Capital Humano) [SFRH/BD/131968/2017]
- ESF (European Social Fund) through PORN (Programa Operacional Regional do Norte) [SFRH/BD/131968/2017]
- Fundação para a Ciência e a Tecnologia [SFRH/BD/131968/2017] Funding Source: FCT
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This study describes the catalytic mechanism of the fungal Thermothelomyces thermophila glucuronoyl esterase (TtGE) using QM/MM methods. Two nearly-degenerate rate-determining transition states were found, and a mutation was proposed to enhance the catalytic rate of TtGE.
Hydrolysis of lignocellulosic biomass, composed of a lignin-carbohydrate-complex (LCC) matrix, is critical for producing bioethanol from glucose. However, current methods for LCC processing require costly and polluting processes. The fungal Thermothelomyces thermophila glucuronoyl esterase (TtGE) is a promising thermophilic enzyme that hydrolyses LCC ester bonds. This study describes the TtGE catalytic mechanism using QM/MM methods. Two nearly-degenerate rate-determining transition states were found, with barriers of 16 and 17 kcal.mol(-1), both with a zwitterionic nature that results from a proton interplay from His346 to either the Ser213-hydroxyl or the lignin leaving group and the rehybridisation of the ester moiety of the substrate to an alkoxide. An oxyanion hole, characteristic of esterases, was provided by the conserved Arg214 through its backbone and sidechain. Our work further suggests that a mutation of Glu267 to a non-negative residue will decrease the energetic barrier in ca. -5 kcal.mol(-1), improving the catalytic rate of TtGE.
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