Synthesis of maleimide-activated carbohydrates as chemoselective tags for site-specific glycosylation of peptides and proteins.

Abstract

An array of maleimide-activated mono- and oligosaccharides were synthesized to permit site-specific glycosylation of cysteine-containing peptides and proteins. Maleimide-activated monosaccharides, in which the native alpha- or beta-O-glycosidic linkages found for nonreducing terminal sugars of native glycoproteins are preserved, were prepared using 2'-aminoethyl glycosides as the key intermediates. In addition, a native high-mannose type oligosaccharide, Man(9)GlcNAc(2)Asn, was converted into its maleimide-activated form by taking advantage of the existing amino group in the Asn portion. The application of these maleimide-activated carbohydrates was exemplified by the site-specific glycosylation of a 36-mer HIV-1 gp41 peptide, T20, which is a potent inhibitor against HIV infection. The chemoselective ligation was found to be rapid, highly efficient, and essentially quantitative. Tagging the biologically active peptide with a mannose and/or oligomannose moiety will be useful for targeting the drug to macrophage and dendritic cells, which are primary targets for HIV-1 infection and are expressing mannose- and oligomanose-specific receptors on their surface. In combination with site-specific mutagenesis, the maleimide-activated carbohydrates can serve as generally applicable tags for site-specific glycosylation of proteins via the highly efficient maleimide-thiol ligation reaction.