Sedimentation equilibrium analysis of five lipocortin-related phospholipase A2 inhibitors from human placenta. Evidence against a mechanistically relevant association between enzyme and inhibitor.
Five proteins from human placenta capable of inhibiting pancreatic phospholipaseA2 were purified. Two of these proteins were identified as lipocortins I and II.The other three proteins were immunologically distinct from lipocortins I and IIand had apparent subunit molecular masses of 32, 33, and 73 kDa as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Amino acid sequence analysis of peptides produced by cyanogen bromide digestion indicated sequence homology of these proteins with lipocortin I and the heavy chain subunit of lipocortin II. Two of these proteins were identified as endonexin II and 67-kDa calelectrin. The third protein appears to be the human form of bovine endonexin I, also characterized as porcine protein II. Sedimentation equilibrium analysis of lipocortin I, endonexin I and II, and the 67-kDa calelectrin suggested monomer-dimer equilibria with dissociation constants in the range of 0.33-1.3 X 10(-3) M and monomer molecular masses of 38,050, 36,400, 36,850, and 73,610 Da, respectively. Self-association of lipocortin II was described by dimerization ofa protomer (K12 = 5.3 x 10(-7) M), followed by an indefinite self-association ofthe dimer (isodesmic dissociation constant