The factor IX phospholipid-binding site is required for calcium-dependent activation of factor IX by factor XIa.
To determine the functional role of the metal-dependent conformational changes in Factor IX, two populations of conformation-specific anti-Factor IX antibodies were prepared. Anti-Factor IX X Mg(II) antibodies bind to Factor IX in the presence of Mg(II) and other metal ions, but not in the absence of metal ions. Anti-Factor IX X Ca(II)-specific antibodies bind to Factor IX in the presence of Ca(II) and Sr(II), but not in the presence of Mn(II), Mg(II), and Ba(II). In the presence of a metal ion that induces the conformational transition recognized by the anti-Factor IX X Mg(II) antibodies, the concentrations of CaCl2 and SrCl2 needed for the half-maximal binding of the anti-Factor IX X Ca(II)-specific antibodies to Factor IX were reduced 3- and 20-fold, respectively. Factor IX binding to phospholipid vesicles was inhibited by the Fab fragments of the anti-Factor IX X Ca(II)-specific antibodies, but was not inhibited by the Fab fragments of the anti-Factor IX X Mg(II) antibodies. Factor XIa activation of Factor IX was also inhibited by the Fab fragments of the anti-Factor IX X Ca(II)-specific antibodies, but not by the anti-Factor IX X Mg(II) antibodies. These results support the hypothesis that Factor IX undergoes two metal-dependent conformational transitions: FIX----FIX'----FIX*. The first transition (FIX----FIX') is metal-dependent but cation-nonselective; the second transition (FIX'----FIX*) is metal-selective for Ca(II) or Sr(II). The second transition results in the expression of conformational determinants necessary for membrane binding and the Ca(II)-dependent activation of Factor IX by Factor XIa. These results suggest chemical similarity between a surface of a domain of Factor XIa and phospholipid vesicles, both of which interact with Factor IX in the presence of Ca(II).
D000906:Antibodies; D000936:Antigen-Antibody Complex; D001665:Binding Sites; D002118:Calcium / Q000494:pharmacology*; D002122:Calcium Chloride / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D004789:Enzyme Activation; D005164:Factor IX / Q000378:metabolism*; D005172:Factor XI / Q000378:metabolism*; D015945:Factor XIa; D006801:Humans; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D015636:Magnesium Chloride; D010743:Phospholipids / Q000097:blood*; D011485:Protein Binding; D011487:Protein Conformation
D000906:Antibodies; D000936:Antigen-Antibody Complex; D010743:Phospholipids; D015636:Magnesium Chloride; D005164:Factor IX; D005172:Factor XI; D004492:Edetic Acid; D015945:Factor XIa; D008274:Magnesium; D002122:Calcium Chloride; D002118:Calcium