Structural Comparison of a Promiscuous and a Highly Specific Sucrose 6F-Phosphate Phosphorylase

Franceus, Jorick; Capra, N.; Desmet, Tom; Thunnissen, A.M.W.H.

doi:10.3390/ijms20163906

Cited by 12 publications

(14 citation statements)

References 52 publications

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“…In the promiscuous SPP from T. thermosaccharolyticum, mutagenesis of Arg134 and His344 causes the activity on fructose 6-phosphate to decrease much more than the activity on fructose, indicating that they may bind the phosphate group of sucrose 6 F -phosphate [8]. Docking experiments of the substrate in the crystal structure of TtSPP later provided further confirmation of this hypothesis [11]. In the strict SPP from I. coccineus, the residues that contribute to the affinity for fructose 6-phosphate, in descending order of importance, are Lys434, Lys364, Arg152 and Tyr377 [11].…”

Section: Enzymementioning

confidence: 91%

“…In this loop, the N(L/V)D(I/L/V)YQ motif that is indicative of SP activity is replaced by a GFDVHQ motif in TtSPP (Figure 2, Figure S1). More recently, phosphorylases from Ruminococcus gnavus E1 and Ilumatobacter coccineus were also found to possess SPP activity [10,11]. Unlike TtSPP, which is very promiscuous and shows high activity on sucrose, these two SPPs are strictly specific to sucrose 6 F -phosphate.…”

Section: Sucrose 6 F -Phosphate Phosphorylasementioning

confidence: 99%

“…The latter SP also shows the highest affinity for sucrose, which may thus be correlated with its stricter specificity in the acceptor binding site. Indeed, the strict SPP from I. coccineus has a much lower K M for its native acceptor fructose 6-phosphate than the promiscuous one from T. thermosaccharolyticum [10,11]. Furthermore, different phosphorylases can exhibit a completely different regioselectivity as well.…”

Section: Transglycosylation For the Synthesis Of Glycosides And Rare mentioning

confidence: 99%

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Sucrose Phosphorylase and Related Enzymes in Glycoside Hydrolase Family 13: Discovery, Application and Engineering

Franceus

Desmet

2020

IJMS

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Sucrose phosphorylases are carbohydrate-active enzymes with outstanding potential for the biocatalytic conversion of common table sugar into products with attractive properties. They belong to the glycoside hydrolase family GH13, where they are found in subfamily 18. In bacteria, these enzymes catalyse the phosphorolysis of sucrose to yield α-glucose 1-phosphate and fructose. However, sucrose phosphorylases can also be applied as versatile transglucosylases for the synthesis of valuable glycosides and sugars because their broad promiscuity allows them to transfer the glucosyl group of sucrose to a diverse collection of compounds other than phosphate. Numerous process and enzyme engineering studies have expanded the range of possible applications of sucrose phosphorylases ever further. Moreover, it has recently been discovered that family GH13 also contains a few novel phosphorylases that are specialised in the phosphorolysis of sucrose 6F-phosphate, glucosylglycerol or glucosylglycerate. In this review, we provide an overview of the progress that has been made in our understanding and exploitation of sucrose phosphorylases and related enzymes over the past ten years.

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Section: Enzymementioning

confidence: 91%

Section: Sucrose 6 F -Phosphate Phosphorylasementioning

confidence: 99%

Section: Transglycosylation For the Synthesis Of Glycosides And Rare mentioning

confidence: 99%

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Sucrose Phosphorylase and Related Enzymes in Glycoside Hydrolase Family 13: Discovery, Application and Engineering

Franceus

Desmet

2020

IJMS

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“…Franceus et al report the characterization of a sucrose 6F-phosphate phosphorylases member (SPP) of the family GH13 subfamily 18 from Ilumatobacter coccineus and its comparison with the enzyme from Thermoanaerobacterium thermosaccharolyticum [15]. Crystal structures of both SPPs were determined to provide insight into their similarities and differences.…”

mentioning

confidence: 99%

“…Considerable differences between the two enzymes were found in the residues responsible for binding the fructose 6-phosphate group in subsite +1. Mutants that provided a higher degree of substrate promiscuity in the SPP from I. coccineus were probed, thus paving the way to rational enzyme engineering in biotechnology [15].…”

mentioning

confidence: 99%

Carbohydrate-Active Enzymes: Structure, Activity, and Reaction Products

Benini¹

2020

IJMS

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Carbohydrate-active enzymes are responsible for both the biosynthesis and breakdown of carbohydrates and glycoconjugates. They are involved in many metabolic pathways, in the biosynthesis and degradation of various biomolecules, such as bacterial exopolysaccharides, starch, cellulose, and lignin, and in the glycosylation of proteins and lipids. Their interesting reactions have attracted the attention of researchers belonging to different scientific fields ranging from basic research to biotechnology. Interest in carbohydrate-active enzymes is due not only to their ability to build and degrade biopolymers, which is highly relevant in biotechnology, but also because they are involved in bacterial biofilm formation, and in the glycosylation of proteins and lipids, which has important health implications.This Special Issue features a collection of research papers and reviews representing an up-to-date state of the art in this growing field of research to broaden our understanding about carbohydrate-active enzymes, their mutants, and their reaction products at the molecular level.Tindara Venuto et al. wrote a paper about alpha2,8-sialyltransferases from ray-finned fish and showed that, in tetrapods, duplicated st8sia genes like st8sia7, st8sia8, and st8sia9 have disappeared while orthologues are maintained in teleosts [1]. They reconstructed the evolutionary history of st8sia genes in fish genomes and by bioinformatics analysis showed changes in the conserved polysialyltransferase domain of the fish sequences possibly accounting for variable enzymatic activities [1].PhNah20A, a β-N-acetylhexosaminidase from the marine bacterium Paraglaciecola hydrolytica S66T, was identified and characterized. By phylogenetic analysis, the authors discovered that PhNah20A is located outside clusters of other studied β-N-acetylhexosaminidases, in a unique position between bacterial and eukaryotic enzymes. PhNah20A is the first characterized member of a distinct subgroup within GH20 β-N-acetylhexosaminidases. The recombinant PhNah20A showed optimum hydrolytic activity on GlcNAc β-1,4 and β-1,3 linkages in chitobiose (GlcNAc)2 and GlcNAc-1,3-β-Gal-1,4-β-Glc (LNT2), the core structure of a human milk oligosaccharide, at pH 6.0 and 50 • C. Interestingly PhNah20A catalyzed the formation of LNT2, the non-reducing trisaccharide β-Gal-1,4-β-Glc-1,1-β-GlcNAc, and in low amounts the β-1,2or β-1,3-linked trisaccharide β-Gal-1,4(β-GlcNAc)-1,x-Glc by a transglycosylation of lactose using 2-methyl-(1,2-dideoxy-α-d-glucopyrano)-oxazoline (NAG-oxazoline) as the donor [2].The production of biofuels from lignocellulosic biomasses is hampered by the saccharification step of the biomasses requiring the concerted action of lignocellulose cellulases and hemicellulases.Zhang et al. characterized the secretome of Trichoderma harzianum EM0925 under induction of lignocellulose cellulases and hemicellulases [3]. Compared with the commercial enzyme preparations, the T. harzianum EM0925 enzyme cocktail presented significantly higher lignocellulolytic enzyme activi...

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Enzymatic Synthesis of Phloretin α‐Glucosides Using a Sucrose Phosphorylase Mutant and its Effect on Solubility, Antioxidant Properties and Skin Absorption

et al. 2021

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Glycosylation of polyphenols may increase their aqueous solubility, stability, bioavailability and pharmacological activity. Herein, we used a mutant of sucrose phosphorylase from Thermoanaerobacterium thermosaccharolyticum engineered to accept large polyphenols (variant TtSPP_R134A) to produce phloretin glucosides. The reaction was performed using 10% (v/v) acetone as cosolvent. The selective formation of a monoglucoside or a diglucoside (53% and 73% maximum conversion percentage, respectively) can be kinetically controlled. MS and 2D‐NMR determined that the monoglucoside was phloretin 4’‐O‐α‐D‐glucopyranoside and the diglucoside phloretin‐4’‐O‐[α‐D‐glucopyranosyl‐(1→3)‐O‐α‐D‐glucopyranoside], a novel compound. The molecular features that determine the specificity of this enzyme for 4’‐OH phenolic group were analysed by induced‐fit docking analysis of each putative derivative, using the crystal structure of TtSPP and changing the mutated residue. The mono‐ and diglucoside were, respectively, 71‐ and 1200‐fold more soluble in water than phloretin at room temperature. The α‐glucosylation decreased the antioxidant capacity of phloretin, measured by DPPH and ABTS assays; however, this loss was moderate and the activity could be recovered upon deglycosylation in vivo. Since phloretin attracts a great interest in dermocosmetic applications, we analyzed the percutaneous absorption of glucosides and the aglycon employing a pig skin model. Although the three compounds were detected in all skin layers (except the fluid receptor), the diglucoside was present mainly on superficial layers.

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Structural Comparison of a Promiscuous and a Highly Specific Sucrose 6F-Phosphate Phosphorylase

Cited by 12 publications

References 52 publications

Sucrose Phosphorylase and Related Enzymes in Glycoside Hydrolase Family 13: Discovery, Application and Engineering

Sucrose Phosphorylase and Related Enzymes in Glycoside Hydrolase Family 13: Discovery, Application and Engineering

Carbohydrate-Active Enzymes: Structure, Activity, and Reaction Products

Enzymatic Synthesis of Phloretin α‐Glucosides Using a Sucrose Phosphorylase Mutant and its Effect on Solubility, Antioxidant Properties and Skin Absorption

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