Redesign of the Active Site of Sucrose Phosphorylase through a Clash-Induced Cascade of Loop Shifts

Sucrose phosphorylases have been applied in the enzymatic production of glycosylated compounds for decades. However, several desirable acceptors, such as flavonoids or stilbenoids, that exhibit diverse antimicrobial, anticarcinogenic or antioxidant properties, remain poor substrates. The Q345F excha...

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Bibliographic Details
Published in:Chembiochem : a European journal of chemical biology 2016-01, Vol.17 (1), p.33-36
Main Authors: Kraus, Michael, Grimm, Clemens, Seibel, Jürgen
Format: Article
Language:English
Subjects:
Bifidobacterium - enzymology
biocatalysis
Catalytic Domain
Crystal structure
Glucosyltransferases - chemistry
Glucosyltransferases - metabolism
Glycosylation
Hydrolysis
Models, Molecular
Molecular Conformation
polyphenol glycosylation
protein engineering
substrate promiscuity
Sucrose
Sucrose - chemistry
Sucrose - metabolism
sucrose phosphorylases
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Summary:Sucrose phosphorylases have been applied in the enzymatic production of glycosylated compounds for decades. However, several desirable acceptors, such as flavonoids or stilbenoids, that exhibit diverse antimicrobial, anticarcinogenic or antioxidant properties, remain poor substrates. The Q345F exchange in sucrose phosphorylase from Bifidobacterium adolescentis allows efficient glucosylation of resveratrol, (+)‐catechin and (−)‐epicatechin in yields of up to 97 % whereas the wild‐type enzyme favours sucrose hydrolysis. Three previously undescribed products are made available. The crystal structure of the variant reveals a widened access channel with a hydrophobic aromatic surface that is likely to contribute to the improved activity towards aromatic acceptors. The generation of this channel can be explained in terms of a cascade of structural changes arising from the Q345F exchange. The observed mechanisms are likely to be relevant for the design of other tailor‐made enzymes. Repositioning key loops by adding a clash‐inducing hydrophobic Phe—remodelling a sucrose phosphorylase into a polyphenol transglucosidase: Introducing a sterically demanding residue into the active site of a sucrose phosphorylase enlarged the active site through indirect effects arising from a cascade of conformational changes. Aromatic acceptors could thus be glucosylated.
ISSN:1439-4227
1439-7633
DOI:10.1002/cbic.201500514