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• W2086532946 abstract "An efficient heterologous expression system for overproduction of the enzyme alpha-1,6-Fucosyltransferase (alpha-1,6-FucT) from Rhizobium sp. has been developed. The gene codifying for the alpha-1,6-FucT was amplified by PCR using specific primers. After purification, the gene was cloned in the plasmid pKK223-3. The resulting plasmid, pKK1,6FucT, was transformed into the E. coli strain XL1-Blue MRF'. The protein was expressed both as inclusion bodies and in soluble form. Changing the induction time a five-fold increase of enzyme expressed in soluble form was obtained. In this way five units of enzyme alpha-1,6-FucT can be obtained per liter of culture. A crude preparation of the recombinant enzyme was used for the synthesis of the branched trisaccharide alpha-D-GlcNAc-(1-->4)-[alpha-L-Fuc-(1-->6)]-D-GlcNAc (3), from chitobiose (2) and GDP-Fucose (1). After purification, the trisaccharide 3 was obtained in a 84% overall yield. In order to elucidate the structural requirements for the acceptors, the specificity of the enzyme was studied towards mono-, di- and trisaccharides, which are structurally related to chitobiose. The enzyme uses, among others, the disaccharide N-acetyl lactosamine as a good substrate; the monosaccharide GlcNAc is a weak acceptor. Finally, several racemic polyhydroxylated indolizidines have been tested as potential inhibitors of the enzyme. Indolizidine 21 was the best inhibitor with an IC50 of 4.5 x 10(-5) M. Interestingly, this compound turned out to be the best mimic for the structural features of the fucose moiety in the presumed transition state." @default.
• W2086532946 created "2016-06-24" @default.
• W2086532946 creator A5007890786 @default.
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• W2086532946 date "2001-06-01" @default.
• W2086532946 modified "2023-10-18" @default.
• W2086532946 title "Heterologous Over-expression ofα-1,6-Fucosyltransferase fromRhizobium sp.: Application to the Synthesis of the Trisaccharideβ-D-GlcNAc(1→4)- [α-L-Fuc-(1→6)]-D-GlcNAc, Study of the Acceptor Specificity and Evaluation of Polyhydroxylated Indolizidines as Inhibitors" @default.
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• W2086532946 doi "https://doi.org/10.1002/1521-3765(20010601)7:11<2390::aid-chem23900>3.0.co;2-0" @default.
• W2086532946 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/11446641" @default.
• W2086532946 hasPublicationYear "2001" @default.
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