Synthesis of Nitrogen-Containing Furanose Sugar Nucleotides for Use as Enzymatic Probes Ryan B. Snitynsky and Todd L. Lowary* Alberta Glycomics Centre and Department of Chemistry, Gunning-Lemieux Chemistry Centre, University of Alberta, Edmonton, Alberta, Canada T6G 2G2 * S Supporting Information ABSTRACT: The sugar nucleotides UDP-2-acetamido-2-deoxy-α-D-galactofuranose (UDP-Galf NAc) and UDP-2-azido-2- deoxy-α-D-galactofuranose (UDP-Galf N 3 ) have been synthesized in preparative scale for the first time. These compounds are useful probes for studying the biosynthesis of glycans containing galactofuranose and/or 2-acetamido-2-deoxy-α-D- galactofuranose residues. S ugar nucleotides are widespread in nature, being the substrates for many glycosyltransferase enzymes. 1 The six- membered ring pyranose forms of sugar nucleotides predom- inate in most biological systems. In contrast, the five-membered ring furanose sugar nucleotides are less common and are absent in mammals. 2 As a result, glycosyltransferases from pathogenic organisms that use furanose sugar nucleotides for cell wall biosynthesis have been proposed as potential drug targets. 2b Campylobacter jejuni, one of the leading causes of bacterial gastroenteritis worldwide, 3 features a 2-acetamido-2-deoxy-α-D- galactofuranose (Galf NAc) moiety within its capsular poly- saccharide (CPS), occasionally modified with a methyl phosphoramidate group at C-3. 4 To understand the pathways that C. jejuni uses to construct its CPS, and potentially study the inhibition of CPS formation, milligram quantities of the sugar nucleotide precursors are required. We describe here the first preparative-scale synthesis of UDP-2-acetamido-2-deoxy-α- D-galactofuranose (UDP-Galf NAc, 1) (Figure 1), the donor species for Galf NAc transferases. The only previously reported preparation of 1 is an enzyme-mediated reaction that interconverts the pyranose and furanose forms of this sugar nucleotide. 5 At equilibrium, the enzymatic reaction produces a 93:7 pyranose/furanose mixture, necessitating a more viable way of producing 1. We also report the synthesis of an azido analogue of 1, UDP-2-azido-2-deoxy-α-D-galactofuranose (UDP-Galf N 3 , 2). We expect 2 to be an efficient probe of both Galf NAc and Galf biosynthesis, similar to how other azido-substituted carbohydrate derivatives have been useful in probing biosynthetic pathways by which complex glycans are assembled. 6 The synthetic precursor to 1, 2-acetamido-2-deoxy-α-D- galactofuranose-1-phosphate (Galf NAc-1-P, 3), was synthe- sized as shown in Scheme 1. 2-Azido-2-deoxy-D-galactopyr- anose (4), obtained through either the azidonitration of 3,4,6- tri-O-acetyl-D-galactal 7 or diazo transfer with galactosamine and TfN 3 , 8 was converted to its furanose form in the presence of 2,2-dimethoxypropane and p-TsOH, 9 followed by deprotection and acetylation to produce 5 in 78% overall yield. Acetolysis yielded a fully acetylated compound 6 in 67% yield, suitable for phosphorylation. Phosphorylation was accomplished by first converting the fully acetylated compound 6 into the corresponding bromide, followed by reaction with dibenzyl phosphate under basic conditions to yield 7 in 61% yield as a 5:1 α/β mixture. For the first step in this transformation, due to the incompatibility of the azido group with HBr, a previously described method 10 employed successfully on the corresponding galactofuranose derivative could not be used. Instead, TiBr 4 was used to form the glycosyl bromide. 11 With the target compound 7 in hand, debenzylation, azide reduction, and amine acetylation were carried out in one step through hydrogenation over Pd/C with acetic anhydride as the solvent. Subsequent removal of the ester protecting groups in a methanol-water-triethylamine solution produced Galf NAc-1-P (3) as a triethylammonium salt in 74% yield. Received: November 6, 2013 Published: December 16, 2013 Figure 1. Structure of UDP-Galf NAc (1, R = NHAc) and UDP- Galf N 3 (2,R=N 3 ). Letter pubs.acs.org/OrgLett © 2013 American Chemical Society 212 dx.doi.org/10.1021/ol4032073 | Org. Lett. 2014, 16, 212-215