Improvement of Fucosylated Oligosaccharides Synthesis by α-L-Fucosidase from Thermotoga maritima in Water-Organic Cosolvent Reaction System.

Mónica A Robles-Arias,Lorena Gómez-Ruiz,Gabriela Rodríguez-Serrano,Salvador R Tello-Solís,Francisco Guzmán-Rodriguez,Sergio Alatorre-Santamaría,Mariano García-Garibay,Alma E Cruz-Guerrero

doi:10.1007/s12010-021-03628-3

Mónica A Robles-Arias, Lorena Gómez-Ruiz + Show 6 more

Open Access

https://doi.org/10.1007/s12010-021-03628-3

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Abstract

The effects of water activity (aw), pH, and temperature on transglycosylation activity of α-L-fucosidase from Thermotoga maritima in the synthesis of fucosylated oligosaccharides were evaluated using different water-organic cosolvent reaction systems. The optimum conditions of transglycosylation reaction were the pH range between 7 and 10 and temperature 90-95°C. The addition of organic cosolvent decreased α-L-fucosidase transglycosylation activity in the following order: acetone > dimethyl sulfoxide (DMSO) > acetonitrile (0.51 > 0.42 > 0.18mM/h). However, the presence of DMSO and acetone enhanced enzyme-catalyzed transglycosylation over hydrolysis as demonstrated by the obtained transglycosylation/hydrolysis rate (rT/H) values of 1.21 and 1.43, respectively. The lowest rT/H was calculated for acetonitrile (0.59), though all cosolvents tested improved the transglycosylation rate in comparison to a control assay (0.39). Overall, the study allowed the production of fucosylated oligosaccharides in water-organic cosolvent reaction media using α-L-fucosidase from T. maritima as biocatalyst.

Highlights

IntroductionHuman milk oligosaccharides (HMOs) are unique to human breast milk (10– 15 g/L)
Highly abundant, human milk oligosaccharides (HMOs) are unique to human breast milk (10– 15 g/L)
In parallel to the pNP-Fuc hydrolysis, some of the released fucose was enzymatically transferred to lactose leading to the formation of fucosylated oligosaccharide while the remaining fucose was observed as free fucose in the reaction medium

Summary

Introduction

Human milk oligosaccharides (HMOs) are unique to human breast milk (10– 15 g/L). In vivo biosynthesis of fucosylated oligosaccharides is catalyzed by specific transferases found in the Golgi apparatus of the mammary alveolar cells [4]. Since these enzymes are difficult to express and purify, as well as require expensive nucleotide-activated substrates, which add to the high cost of their production, their usage for the in vitro synthesis of fucosylated oligosaccharides is limited [5, 6]. The most recent reports have described successful protocols to obtain fucosylated oligosaccharides in the α-L-fucosidase assisted enzymatic reactions [9,10,11,12]

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