Development of an in vivo glucosylation platform by coupling production to growth: Production of phenolic glucosides by a glycosyltransferase of Vitis vinifera.
Glycosylation of small molecules can significantly alter their properties such as solubility, stability, and/or bioactivity, making glycosides attractive and highly demanded compounds. Consequently, many biotechnological glycosylation approaches have been developed, with enzymatic synthesis and whole-cell biocatalysis as the most prominent techniques. However, most processes still suffer from low yields, production rates and inefficient UDP-sugar formation. To this end, a novel metabolic engineering strategy is presented for the in vivo glucosylation of small molecules in Escherichia coli W. This strategy focuses on the introduction of an alternative sucrose metabolism using sucrose phosphorylase for the direct and efficient generation of glucose 1-phosphate as precursor for UDP-glucose formation and fructose, which serves as a carbon source for growth. By targeted gene deletions, a split metabolism is created whereby glucose 1-phosphate is rerouted from the glycolysis to product formation (i.e., glucosylation). Further, the production pathway was enhanced by increasing and preserving the intracellular UDP-glucose pool. Expression of a versatile glucosyltransferase from Vitis vinifera (VvGT2) enabled the strain to efficiently produce 14 glucose esters of various hydroxycinnamates and hydroxybenzoates with conversion yields up to 100%. To our knowledge, this fast growing (and simultaneously producing) E. coli mutant is the first versatile host described for the glucosylation of phenolic acids in a fermentative way using only sucrose as a cheap and sustainable carbon source.
Centre of Expertise - Industrial Biotechnology and Biocatalysis, Department of Biochemical and Microbial Technology, Ghent University, Coupure links 653, 9000, Ghent, Belgium. Frederik.DeBruyn@Ugent.be.
Frederik
F
De Bruyn
Centre of Expertise - Industrial Biotechnology and Biocatalysis, Department of Biochemical and Microbial Technology, Ghent University, Coupure links 653, 9000, Ghent, Belgium.
Brecht
B
De Paepe
Centre of Expertise - Industrial Biotechnology and Biocatalysis, Department of Biochemical and Microbial Technology, Ghent University, Coupure links 653, 9000, Ghent, Belgium.
Jo
J
Maertens
Centre of Expertise - Industrial Biotechnology and Biocatalysis, Department of Biochemical and Microbial Technology, Ghent University, Coupure links 653, 9000, Ghent, Belgium.
Joeri
J
Beauprez
Centre of Expertise - Industrial Biotechnology and Biocatalysis, Department of Biochemical and Microbial Technology, Ghent University, Coupure links 653, 9000, Ghent, Belgium.
Pieter
P
De Cocker
Research Group SynBioC, Department of Sustainable Organic Chemistry and Technology, Faculty of Bioscience Engineering, Ghent University, Coupure links 653, 9000, Ghent, Belgium.
Stein
S
Mincke
Research Group SynBioC, Department of Sustainable Organic Chemistry and Technology, Faculty of Bioscience Engineering, Ghent University, Coupure links 653, 9000, Ghent, Belgium.
Christian
C
Stevens
Centre of Expertise - Industrial Biotechnology and Biocatalysis, Department of Biochemical and Microbial Technology, Ghent University, Coupure links 653, 9000, Ghent, Belgium.
Marjan
M
De Mey
D004734:Energy Metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005632:Fructose / Q000378:metabolism; D017353:Gene Deletion; D005958:Glucosephosphates / Q000378:metabolism; D005960:Glucosides / Q000378:metabolism*; D005964:Glucosyltransferases / Q000235:genetics / Q000378:metabolism; D016695:Glycosyltransferases / Q000235:genetics / Q000378:metabolism*; D060847:Metabolic Engineering / Q000379:methods*; D030781:Organisms, Genetically Modified / Q000235:genetics / Q000378:metabolism; D010636:Phenols / Q000378:metabolism*; D014532:Uridine Diphosphate Glucose / Q000378:metabolism; D027843:Vitis / Q000201:enzymology* / Q000235:genetics
D005958:Glucosephosphates; D005960:Glucosides; D010636:Phenols; D005632:Fructose; C031590:glucose-1-phosphate; D016695:Glycosyltransferases; D005964:Glucosyltransferases; C025901:sucrose phosphorylase; D014532:Uridine Diphosphate Glucose
Escherichia coli W; UDP-glucose; glucogallin; glucosylation; metabolic engineering; phenolic acid