Chemoenzymatic synthesis of thio-nod factor intermediates — Enzymatic transfer of glucosamine on thiochitobiose derivatives

Abstract

The chemoenzymatic syntheses of thioanalogues of nodulation factors in which the nonreducing end glucosamine residue is available for the introduction of the fatty acid moiety at the free NH2 group are reported. We are describing the chemical synthesis of UDP-GlcNH2 and its use in the enzymatic transfer of GlcNH2 by the bovine galactosyltransferase (EC 2.4.1.90) onto O-4 of the nonreducing end N-acetylglucosamine residues of chitobiose, thiochitobiose, and allyl thiochitobioside. The enzymatic reactions on chitobiose and thiochitobiose were followed by TLC and MALDI MS and showed about 50% conversion of the disaccharides to the desired products. However, these reducing trisaccharides could not be obtained totally free of salts and degraded on ion exchange chromatography. Thus, we investigated the enzymatic transfer on the nonreducing allyl thiochitobioside analogue. We describe here the chemical synthesis of this thiodisaccharide and the enzymatic transfer of GlcNH2 at O-4 of its nonreducing end glucosamine residue to give the desired allyl thiotrisaccharide. This thiotrisaccharide was obtained pure in 41% yield and was characterized by 1H NMR (HSQC) and HRMS.Key words: nodulation factors, synthesis, enzymatic transfer, thiooligosaccharides, UDP-glucosamine.

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