Characterisation of a hyperthermophilic transketolase from Thermotoga maritima DSM3109 as a biocatalyst for 7-keto-octuronic acid synthesis.

Max Cárdenas-Fernández,John M Ward,Dragana Dobrijevic,Fabiana Subrizi,Helen C Hailes

doi:10.1039/d1ob01237a

Max Cárdenas-Fernández, John M Ward + Show 3 more

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https://doi.org/10.1039/d1ob01237a

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Abstract

Transketolase (TK) is a fundamentally important enzyme in industrial biocatalysis which carries out a stereospecific carbon–carbon bond formation, and is widely used in the synthesis of prochiral ketones. This study describes the biochemical and molecular characterisation of a novel and unusual hyperthermophilic TK from Thermotoga maritima DSM3109 (TKtmar). TKtmar has a low protein sequence homology compared to the already described TKs, with key amino acid residues in the active site highly conserved. TKtmar has a very high optimum temperature (>90 °C) and shows pronounced stability at high temperature (e.g. t1/2 99 and 9.3 h at 50 and 80 °C, respectively) and in presence of organic solvents commonly used in industry (DMSO, acetonitrile and methanol). Substrate screening showed activity towards several monosaccharides and aliphatic aldehydes. In addition, for the first time, TK specificity towards uronic acids was achieved with TKtmar catalysing the efficient conversion of d-galacturonic acid and lithium hydroxypyruvate into 7-keto-octuronic acid, a very rare C8 uronic acid, in high yields (98%, 49 mM).

Highlights

Biocatalysis is considered as a green and sustainable technology based on the principles and metrics of green chemistry and sustainable development.[1]
These enzymes possess an optimum temperature above 50 °C and extended activity half-life (t1/2) at high temperatures, being more robust under demanding industrial applications such as high temperatures and the presence of organic solvents compared to their mesophilic counterparts.[20,21]
It has a low protein homology related to well-known TKs, with 10% of amino acid residues highly conserved especially the ones in the active site

Summary

Introduction

Biocatalysis is considered as a green and sustainable technology based on the principles and metrics of green chemistry and sustainable development.[1]. TKtmar optima temperature and pH were determined by measuring the initial rates (for up to 20 min) of the reaction between glycolaldehyden (GA) and LiHPA to yield L-erythrulose (Ery).

Results

Conclusion