Yuko Nakahara

Solid-phase synthesis of an O-linked glycopeptide based on a benzyl-protected glycan approach

The solid-phase synthesis of asialo-[Ala18]-B-chain (2) of human alpha 2HS glycoprotein is described. Disaccharide-linked serine unit 12, carrying a benzyl protecting group, was synthesized via stereoselective glycosylation of 8 with 6. Peptide synthesis was carried out by the Fmoc method utilizing an automated peptide synthesizer. A modified procedure using a mechanical shaker at the coupling step with 12 made easy the recovery of unreacted 12. The benzylated glycopeptide thus synthesized was cleaved from the resin and hydrogenated to give 2.

Synthesis of a glycopeptide carrying a N-linked core pentasaccharide.

A glycopeptide carrying a pentasaccharide core structure of asparagine-linked glycoproteins was synthesized. The synthesis of the carbohydrate part was performed starting from monosaccharide components in an unambiguous manner. The resultant pentaglycosyl azide was reduced into corresponding glycosyl amine and coupled with an aspartic acid derivative to furnish an Asn-linked oligosaccharide in a protected form. Subsequent coupling with a dipeptide, followed by deprotection gave the target compound.

Efficient sequential segment coupling using N-alkylcysteine-assisted thioesterification for glycopeptide dendrimer synthesis.

A highly pure MUC1-derived glycopeptide dendrimer of 22 kDa was prepared by a sequential segment coupling, achieved by an N-alkylcysteine (NAC)-assisted thioesterification. The glycopeptide having C-terminal NAC was prepared by the Fmoc method and converted to the thioester by 3-mercaptopropionic acid treatment. The thioester was condensed with a lysine trimer carrying NAC to afford tetramer, which was then converted to the thioester. Two tetramers were condensed with ethylenediamine to give the octameric glycopeptide dendrimer.

Chemoenzymatic synthesis of hydrophobic glycoprotein: synthesis of saposin C carrying complex-type carbohydrate.

The complex-type N-linked octasaccharide oxazoline having LacNAc as the nonreducing end sugar was efficiently synthesized using the benzyl-protected LacNAc, mannose, and β-mannosyl GlcNAc units as key building blocks. To achieve a highly β-selective glycosylation with the LacNAc unit, the N-trichloroacetyl group was used for the protection of the amino group in the LacNAc unit. After complete assembly of these units and deprotection, the obtained free sugar was successfully derivatized into the corresponding sugar oxazoline. On the other hand, the N-acetylglucosaminylated saposin C, a hydrophobic lipid-binding protein, was chemically synthesized by the native chemical ligation reaction. On the basis of the previous results related to the synthesis of the nonglycosylated saposin C, the O-acyl isopeptide structure was introduced to the N-terminal peptide thioester carrying GlcNAc to improve its solubility toward aqueous organic solvents. The ligation reaction efficiently proceeded with the simultaneous O- to N-acyl shift at the O-acyl isopeptide moiety. After the removal of the cysteine-protecting group and folding, saposin C carrying GlcNAc was successfully obtained. The synthetic sugar oxazoline was then transferred to this glycoprotein using the mutant of endo-β-N-acetylglucosaminidase from Mucor hiemalis (Endo-M) (glycosynthase), and the saposin C carrying the complex-type nonasaccharide was successfully obtained.

Solid-phase synthesis of CD52 glycopeptide and an efficient route to Asn-core pentasaccharide conjugate

The intact peptide sequence (18) as well as its glycoform carrying an N-linked core pentasaccharide (1) of CD52 antigen were prepared by means of solid-phase synthesis employing Fmoc-amino acids and benzyl-protected oligosaccharide-asparagine conjugate (3) as building blocks. It was concluded that the pentasaccharide structure had little influence on further peptide elongation in solid-phase synthesis and the benzylated pentasaccharide moiety was sufficiently stable to the 95% TFA acidic conditions used to release glycopeptide from the supporting resin. The paper also describes an efficient route leading to asparagine-core pentasaccharide conjugate (3) which was prepared in seven steps for an overall yield of 23% from monosaccharide units 5, 6, 7 and 8.

Rationally designed syntheses of high-mannose and complex type undecasaccharides

Synthetic routes are described to a high-mannose type triantenary undecasaccharide 1 and a completely protected form 39 of the complex type biantenary undecasaccharide 2 carrying alpha-(2-->3)-linked sialic acid residues. Starting from a previously reported trisaccharide 4, the core pentasaccharides 15 and 37 were synthesized through regio- and/or stereo-selective mannosylation with a suitably protected mannosyl donor. Chain elongation of 15 by stepwise addition of a mannose residue afforded an undecasaccharide 20 that was eventually deprotected to give 1. On the other hand, coupling of 38 and a trisaccharide glycosyl donor 36 afforded 39.

Chemoselective fluorination for primary alcohols

Abstract Primary alcohols and their silylated derivatives are selectively fluorinated by tetraalkylammonium fluoride and aryl (or alkyl )sulfonyl fluoride.

Solid-phase synthesis of the B-chain of human α2HS glycoprotein

Abstract The B-chain of human α2HS glycoprotein 1, a heptacosapeptide carrying a trisaccharide (sialyl T) side chain, was synthesized. Prior to the Fmoc-based solid-phase synthesis of the glycopeptide, the benzyl-protected glycosyl serine building block 6 was prepared via β-stereoselective glycosylation of the 2-azido-2-deoxygalactosyl serine 11 with the sialyl galactosyl trichloroacetimidate 9. An automated peptide synthesizer was efficiently used for the elongation of the entire peptide chain except for the coupling with 6. The synthesized glycopeptide was cleaved from the resin by the TFA method. The resultant mixture of the benzylated glycopeptides was treated with TMSOTf–thioanisole in TFA and then with aq NaHCO3 and 1,4-dithiothreitol to give 1.

Synthesis of the root nodule-inducing factor NodRm-IV(C16:2,S) of rhizobium meliloti and related compounds☆

Abstract Nod factors NodRm-IV(RCO,S) that carry natural as well as unnatural fatty acids were synthesized in a stereocontrolled manner.

Chemical synthesis of mouse pro-opiomelanocortin(1–74) by azido-protected glycopeptide ligation via the thioester method

The thioester method is a peptide condensation reaction, which requires the protection of Lys side chains for chemoselective ligation. We recently found that the azido group could be used as an amino protecting group in the peptide condensation by the thioester method. In this study, we synthesized the glycosylated mouse pro-opiomelanocortin (1-74) by the thioester method. The N-terminal peptide thioester segment, whose Lys side chain was protected by an azido group, was prepared using a 9-fluorenylmethoxycarbonyl (Fmoc) strategy and an N-alkylcysteine (NAC)-assisted thioesterification reaction. The C-terminal azido-glycopeptide segment carrying N- and O-linked glycans was also prepared by the Fmoc chemistry and condensed with the N-terminal segment by the silver ion-free thioester method. These results showed that our azido-based strategy was fully compatible with the NAC-assisted method and glycoprotein synthesis.