Limulin: a C-reactive protein from Limulus polyphemus.

A protein which binds specifically to the phosphorylcholine residues of a phosphorylcholine affinity column in the presence of CaZ+ has been isolated from the hemolymph of the horseshoe crab Limuluspolyphemus. Immunological cross-reactivity of the phosphorylcholine-binding protein with limulin, a sialic acid-specific lectin also found in the hemolymph prepared by a different method, was shown by the formation of a single line of identity on immunodiffusion plates using antisera prepared against the phosphorylcholine-binding protein. The Limulus C-reactive protein (CRP) isolated by the phosphorylcholine affinity column precipitates with the C-polysaccharide of pneumococcus and with a synthetic derivative of bovine serum albumin to which phosphorylcholine is covalently attached. Precipitation is inhibited by either EDTA or by phosphorylcholine. This protein also agglutinates horse red blood cells and shows weak cross-reactivity with sheep antisera prepared against rabbit C-reactive protein. The hemolymph hemagglutination titer is markedly decreased by pretreatment of the hemolymph with antisera prepared against the Limulus phosphorylcholine-binding protein. Phosphorylcholine does not inhibit the hemagglutination by whole hemolymph or by Limulus phosphorylcholine-binding protein, but a protein containing sialic acid oligosaccharides does inhibit the hemagglutination. Electron spin resonance experiments using a spin label which resembles phosphorylcholine shows binding of the spin label to the protein only in the presence of Ca”. M&+ cannot substitute for Ca2+ in supporting the binding of spin-labeled phosphorylcholine to limulin. The spin label can be disassociated from the protein by EDTA or competitively removed by phosphorylcholine, but not by phosphate ions or by choline. The relationship of limulin to the C-reactive proteins of rabbit and man are discussed.