Hot couplings without racemization: Protected peptides featuring C‐terminal pseudoprolines were synthesized on a solid support, and these versatile building blocks were used in convergent peptide‐segment couplings, which proceeded without racemization even under microwave conditions. The solubility‐enhancing effect of pseudoproline residues facilitated the synthesis of complex RNase 39‐mer glycopeptide thioesters.
Recombinant human erythropoietin (EPO) is the main therapeutic glycoprotein for the treatment of anemia in cancer and kidney patients.T he in-vivo activity of EPO is carbohydrate-dependent with the number of sialic acid residues regulating its circulatory half-life.E PO carries three Nglycans and thus obtaining pure glycoforms provides am ajor challenge.W eh ave developed ar obust and reproducible chemoenzymatic approach to glycoforms of EPO with and without sialic acids.E PO was assembled by sequential native chemical ligation of two peptide and three glycopeptide segments.The glycopeptides were obtained by pseudoprolineassisted Lansbury aspartylation. Enzymatic introduction of the sialic acids was readily accomplished at the level of the glycopeptide segments but even more efficiently on the refolded glycoprotein. Biological recognition of the synthetic EPOs was shown by formation of 1:1c omplexes with recombinant EPO receptor.
A convergent synthesis for erythropoietin (EPO) 1-28 N-glycopeptide hydrazides was developed. In this approach, EPO 1-28 peptides were synthesized on the solid phase and converted to C-terminal hydrazides after cleavage from the resin. After selective deprotection of the Asp24 side chain, the desired glycosylamine was coupled by pseudoproline-assisted Lansbury aspartylation. Although the initial yields of the EPO 1-28 glycopeptides were satisfactory, they could be markedly improved by increasing the purity of the peptide using a reversed-phase high-performance liquid chromatography (RP-HPLC) purification of the protected peptide.
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