Studies on Limulus amoebocyte lysate. Isolation of pro-clotting enzyme.
A pro-clotting enzyme capable of causing the gelation of clottable proteins in Limulus polyphemus (horseshoe crab) has been purified to apparent homogeneity as judged by sodium dodecyl sulfate-gel electrophoresis. The activation of the pro-clotting enzyme depended on the presence of both Ca+ and endotoxin. It contained gamma-carboxyglutamic acids and gave a single NH2-terminal lysine. The enzyme was inhibited by diisopropyl fluorophosphate, phenylmethylsulfonyl fluoride, and soy bean trypsin inhibitor, indicating that it is a serine protease. The molecular weight of the proclotting enzyme was determined to be at least 150,000 by sodium dodecyl sulfate-gel electrophoresis under reducing and denaturing conditions. The protein appears to consist of a single peptide chain, since exposure of the reduced and carboxymethylated enzyme to 6 M guanidine hydrochloride failed to dissociate it into any subunits.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001181:Arthropods / Q000201:enzymology*; D001779:Blood Coagulation Factors / Q000302:isolation & purification* / Q000378:metabolism; D010450:Endopeptidases; D004731:Endotoxins / Q000032:analysis*; D004792:Enzyme Precursors / Q000302:isolation & purification / Q000378:metabolism; D005782:Gels; D006737:Horseshoe Crabs / Q000201:enzymology*; D007711:Klebsiella pneumoniae
D000596:Amino Acids; D001779:Blood Coagulation Factors; D004731:Endotoxins; D004792:Enzyme Precursors; D005782:Gels; D010450:Endopeptidases; C014775:pro-clotting enzyme