Synthesis and application of sialic acid-containing building blocks for glycopeptide libraries. Establishing glycosylation conditions

Three different sialic acid-containing building blocks (6-8) were synthesized for use in solid-phase glycopeptide libraries. Investigation of the conditions for glycosylation of threonine (Thr) with various sialic acid donors revealed that the best results were obtained by coupling glycosyl xanthate 2 to the acceptors Fmoc-Thr-OH (5) or the alpha-azido acid analogue of Thr, 4. Among several catalysts employed, phenylsulfanyl triflate (PST) afforded the best yields. Both the N-Fmoc and alpha-azido analogues of Thr allowed glycosylation with good stereoselectivity in 80% (--> 8) and 84% (--> 6) yield, respectively. Introduction of a phenylthio group in the 3 position of the sialic acid donor 3, to assist the stereoselective outcome of the glycosylation reaction, gave good results; however difficulties in the removal of the phenylthio auxiliary group made this route less attractive. Both building blocks 6 and 8 were successfully introduced in solid-phase glycopeptide synthesis. Interestingly, alkaline deprotection of the Fmoc group of 8, necessary for subsequent introduction of amino acids, resulted in an immediate attack of the alpha-amino group on the sialic acid methyl ester to form the lactam 14. This side reaction was also observed during reduction of the azido acid building block 6 under alkaline conditions, but could be suppressed by performing the reduction under acidic conditions. Lactam formation was completely avoided by hydrolysis of the methyl ester prior to reduction of the azide.