INVOLVEMENT OF ALPHA-2-MACROGLOBULIN AND C-REACTIVE PROTEIN IN A COMPLEMENT-LIKE HEMOLYTIC SYSTEM IN THE ARTHROPOD, LIMULUS-POLYPHEMUS

Homologues of two plasma proteins of vertebrates, alpha2-macroglobulin and C-reactive protein, participate in a hemolytic system of the ancient arthropod, Limulus polyphemus. C-reactive protein, which can under the appropriate circumstances activate the classical pathway of the mammalian complement system, is an essential element of the hemolytic system of Limulus. The selective removal of C-reactive protein from the plasma with phosphorylethanolamine-agarose inactivated hemolysis. Addition of affinity-purified C-reactive protein to inactive plasma restored activity. Exposure of plasma to phosphorylethanolamine in solution potentiated hemolysis. Alpha2-macroglobulin is a member of the same protein family as the complement protein C3 and both require an intact thiol ester for activity. Treatment of Limulus plasma with methylamine under conditions that inactivate thiol-ester-containing proteins reduced the hemolytic activity of some plasma preparations. Addition of purified Limulus alpha2-macroglobulin to the methylamine-treated plasma restored hemolytic activity. However, alpha2-macroglobulin is not necessary for hemolysis since the hemolytic activity of some pooled plasma preparations was insensitive to methylamine treatment under conditions that inactivated alpha2-macroglobulin. Purified C-reactive protein was hemolytic in the absence of alpha2-macroglobulin. These observations suggest that the proteins in Limulus plasma that participate in hemolysis represent the components of an ancient invertebrate defense system with distant evolutionarily affinities to the vertebrate complement system.