Two forms of factor C from the amoebocytes of Carcinoscorpius rotundicauda: purification and characterisation.
The two apparent form of the endotoxin-sensitive Factor C which were found to exist in the amoebocytes of horseshoe crabs have been separately purified to homogeneity from the lysate of the South-East Asian species, Carcinoscorpius rotundicauda. Both forms are serine proteinase zymogens having an apparent molecular mass of 132 kDa. By reducing SDS-PAGE, one was shown to consist of a single polypeptide while the other has a heavy chain (80 kDa) and a light chain (52 kDa) bridged by disulfide linkage(s). Both zymogen forms have endotoxin (lipopolysaccharide) receptors to which endotoxin binds to activate their catalytic sites. However, single-chain Factor C appears to have higher-affinity endotoxin-binding sites which are competitively but reversibly occupied by DMSO when the latter was added during its purification. Another salient difference between the two forms of Factor C is exhibited in their manner of activation by endotoxin. While double-chain Factor C appears similar to that of Tachypleus tridentatus, single-chain Factor C did not undergo any proteolytic cleavage upon activation. This conformational transition of zymogen activation suggests an alternative reversible pathway of endotoxin activation for the single-chain Factor C.
Department of Zoology, National University of Singapore.
J L
JL
Ding
M A
MA
Navas
B
B
Ho
D000818:Animals; D060829:Arthropod Proteins; D001665:Binding Sites; D001667:Binding, Competitive; D004121:Dimethyl Sulfoxide / Q000737:chemistry; D004731:Endotoxins / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D004792:Enzyme Precursors / Q000302:isolation & purification*; D006434:Hemocytes / Q000737:chemistry*; D006737:Horseshoe Crabs / Q000737:chemistry*; D007527:Isoenzymes / Q000302:isolation & purification*; D012697:Serine Endopeptidases / Q000302:isolation & purification*
D060829:Arthropod Proteins; D004731:Endotoxins; D004792:Enzyme Precursors; D007527:Isoenzymes; D012697:Serine Endopeptidases; C057151:Limulus clotting factor C; D004121:Dimethyl Sulfoxide