Interferon-gamma(IFN-y)is an important cytokine with enhanced immune activity,anti-tumor and antiviral ef-fects,and holds significant potential in medical research and clinical applications.In this study,we report a highly efficient semi-synthesis strategy of homogeneous N-GlcNAc modified IFN-y.The glycopeptide fragment(Pyr1-Leu33)and peptide fragment(Lys34-Ser39)were prepared through chemical methods.And the peptide fragment(Ser40-Gly138)was obtained through Escherichia coli(E.coli)expression.Subsequently,using a combination of"expressed serine ligation"and"native chemical ligation-desulfurization",we ligated these fragments from the C-terminal to the N-terminal,resulting in a full-length glycoprotein,which was successfully refolded to obtain the desired product.
native chemical ligationexpressed serine ligationβ-thioamino acidglycosylation
NETL
NSTL
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