Biological activity, membrane-targeting modification, and crystallization of soluble human decay accelerating factor expressed inE. coli

摘要: Decay-accelerating factor (DAF, CD55) is a glycophosphatidyl inositol-anchored glycoprotein that regulates the activity of C3 and C5 convertases. In addition to understanding mechanism complement inhibition by DAF through structural studies, there also an interest in possible therapeutic potential molecule. this report we describe cloning, expression Escherichia coli, isolation membrane-targeting modification four short consensus repeat domains soluble human with additional C-terminal cysteine residue permit site-specific modification. The purified refolded recombinant protein was active against both classical alternative pathway assays activation had similar biological expressed Pichia pastoris. Modification membrane-localizing peptide restored cell binding gave large increase antihemolytic potency. These data suggested correctly folded suitable for studies as well being basis DAF-derived therapeutic. Crystals E. coli-derived were obtained diffracted 2.2 A, thus permitting first detailed X-ray crystallography on functionally regulator direct potential.