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FVIII is a protein cofactor essential for blood coagulation. The deficiency of factor VIII activity in humans is associated with a congenital bleeding disorder, called hemophilia A.1) In general, most of chemical reaction such as surface modification of a protein is performed at a high pH. In contrast Factor VIII should be performed at pH 6.5~7.5 to prevent the lees of activity.2) In order to enhance, Factor VIII’s activity, usually thiolation performed at pH 7.5. The purpose of Factor VIII thiolation enhanced PEGylation yield. Before Thiolated Factor VIII PEGylation, human IgG
selected as a model-protein for thiol PEGylation. Because IgG(150kDa) has similar molecular weight compared with rFactor VIII(170kDa). Thiolation of human IgG performed with N-succinimidyl S-acetylthioacetate (SATA), and then amino residue of human IgG was
labeledby su lfhydryl. After that SATA-labelede human IgG performed with hydroxylamine, and then human IgG has more sulfide residue. Free N-succinimidyl S-acetylthioacetate (SATA) was separated using Sephadex G-25 column. Thiolated IgG PEGylation was performed with PEG-SH
(MW 30kDa). And analysis of thiol PEGylation, used by HPLC, GPC and MALDI-TOF.