Effects of Ca2+ ions on the horseshoe crab coagulation cascade triggered by lipopolysaccharide

The lipopolysaccharide (LPS)-triggered horseshoe crab coagulation cascade is composed of three protease zymogens, prochelicerase C (proC), prochelicerase B (proB) and the proclotting enzyme (proCE). In this study, we found that Ca (2+) ions increase the production of the clotting enzyme as a result of a cascade reaction reconstituted by recombinant proteins of wild-type (WT) proC, WT proB and WT proCE. We divided the cascade into three stages: autocatalytic activation of WT proC on the surface of LPS into WT alpha-chelicerase C (Stage 1); activation of WT proB on the surface of LPS into WT chelicerase B by WT alpha-chelicerase C (Stage 2) and activation of WT proce into WT CE by chelicerase B (Stage 3). Ca2+ ions enhanced the proteolytic activation in Stage 2, but not those in Stages 1 and 3. Moreover, we performed isothermal titration calorimetry to clarify the interaction of LPS or the recombinant zymogens with Ca2+ ions. LPS interacted with Ca2+ ions at an association constant of K-a = 4.7 x 10(4) M-1, but not with any of the recombinant zymogens. We concluded that LPS bound with Ca2+ ions facilitates the chain reaction of the cascade as a more efficient scaffold than LPS itself.

Automated summary

Through experiments, it was found that calcium ions can increase the production of clotting enzymes. The coagulation cascade, consisting of three protease zymogens with different functions, can be divided into three stages, and calcium ions only enhance protein activation in the second stage. At the same time, it was confirmed that lipopolysaccharide can interact with calcium ions to facilitate the occurrence of the cascade reaction.