Recombinant Factor C from Carcinoscorpius Rotundicauda Binds Endotoxin
The coagulation cascade in the circulating amoebocytes of the Singapore horseshoe crab, Carcinoscorpius rotundicauda, plays a vital role in the crab’s defence against invading Gram negative bacteria present in its habitat. Factor C, a serine protease zymogen, is activated by Gram negative bacterial endotoxin or lipopolysaccharide (LPS) to initiate this coagulation cascade. The Limulus gelation assay for detection of endotoxin is based on the affinity of the endotoxin-binding domain of Factor C for LPS [Ho, 1983; Levin, 1985]. Consequently, C. rotundicauda Factor C cDNA has been cloned and expressed in the methylotrophic yeast, Pichia pastoris, to produce recombinant Factor C (rFC) [Roopashree et al., 1996] which will serve as an alternative source of the enzyme to the conventional amoebocyte lysate. This paper describes the biological activity of full-length rFC viz its immunoreactivity by Western detection and particularly, the functionality of its endotoxinbinding domain to bind specifically to LPS and lipid A.
KeywordsPichia Pastoris Coagulation Cascade Methylotrophic Yeast Yeast Lysate Yeast Cell Lysate
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