Method of preparing protein conjugate

A method for preparing a protein conjugate, the method comprising preparing a conjugate by linking a mono-PEGylated immunoglobulin Fc region, which is prepared by linking a linker of Formula 1 below to the N-terminus of an immunoglobulin Fc region comprising a hinge sequence, to C-terminal cysteine of a glucagon-like peptide-1 (GLP-1) receptor/glucagon receptor dual agonist comprising the amino acid sequence set forth in SEQ ID NO:

• 1 or 2:
• CHO-L1-(OCH2CH2)nO-L2-R [Formula 1]
• wherein in Formula 1:
• L1 is a straight- or branched-chain C1-C6 alkylene;
• L2 is -a1-CONH-, -a1-NHCO-, -a1-NHCO-a2-, -COO-, -b1-COO-, -COO-b2-, or -b1-COO-b2-, and a1, a2, b1, and b2 are each independently a straight- or branched-chain C1-C6 alkylene;
• n is from 10 to 2,400;
• and
• R is any one selected from the group consisting of 2,5-dioxopyrrolidinyl, 2,5-dioxopyrrolyl, aldehyde, maleimide, C6-C20 aryl disulfide, C5-C20 heteroaryl disulfide, vinyl sulfone, thiol, halogenated acetamide, succinimide, p-nitrophenyl carbonate, thioester, and derivatives thereof.