Synthesis Of Di-thiolated O-antigen Of Escherichia Coli Serotypes 64 And Fragment Of Zwitterionic Polysaccharides A2

Escherichia coli（E.coli）is a Gram-negative bacterium and one of the three"critical priority"antibiotic-resistant pathogens listed by the World Health Organization in 2017.Pathogenic E.coli can cause a range of infections,from gastroenteritis to extraintestinal infections such as urinary tract,bloodstream,and central nervous system infections.There is an urgent need to develop glycoconjugate vaccines based on E.coli surface polysaccharides.Common glycoantigens are often easily degraded by enzymes in the body.In order to enhance the tolerance of common glycoantigens,we focused on the E.coli surface lipopolysaccharide serotype 64 O-antigen and successfully completed the first total synthesis of the di-thiolated O-antigen pentasaccharide.This O-antigen has a branched pentasaccharide repeat unit with the structure:→3)-D-Glc A-β-（1→3）-[D-Galp-β-（1→6）-D-Galp-β-（1→3）]-D-Glcp NAc-β-（1→3）-D-Manp NAc-α-(1→.The molecule has a less active glucuronic acid and a unique 1,3-di-thiolated structure.We synthesized the glucuronic acid donor 2-9 through pre-glycosylation oxidation,and obtained the branched trisaccharide 2-52 with an anomeric C1-STol structure through linear synthesis strategy[1+（1+1）],which has the potential to efficiently convert to C1-β-SH.The assembly of three key glycosyl fragments,trisaccharide thiol 2-43,3-O-acetyl-2-nitroglucal 2-5,and monosaccharide thiol 2-73,was achieved using the"one-pot relay S-glycosylation"synthetic method developed by our research group.A one-pot with two-steps procedure successfully constructedβ-（1→3）-di-thiolated glycosidic bond.In addition,zwitterionic polysaccharides（ZPS）have unique immunological properties that can directly activate T cell-dependent immune responses.The synthesis of zwitterionic polysaccharide PS A2 has not been reported yet.In order to further reveal the immune mechanism of ZPS and apply its unique immunogenicity to the development of glycoconjugate vaccines,we designed and synthesized the important glycosyl fragment 3-5 of the zwitterionic polysaccharide PS A2 O-antigen.During the introduction of 2-N₃,we optimized the reaction conditions about the safer azide reagent TMSN₃（trimethylsilyl azide）instead of Na N₃ and successfully achieved efficient conversion from D-glucose to 2-azido-2-deoxy-D-mannose.By introducing TCA（trichloroacetyl）protection to the amino group of the monosaccharide fragment in advance and using its neighboring group to participate in the glycosylation reaction,the disaccharide product 3-28 was obtained with excellent stereoselectivity.