Synthetic factor VIII peptides with amino acid sequences contained within the C2 domain of factor VIII inhibit factor VIII binding to phosphatidylserine.
The effective activation of factor X by factor IXa requires the co-factor activity of activated factor VIII (FVIII). Factor Xa formation is also dependent on the presence of negatively charged phospholipid. A phospholipid binding domain of FVIII has been reported to be present on the FVIII light chain. Recent observations on a subset of human FVIII inhibitors have implicated the carboxyl-terminal C2 domain of FVIII as containing a possible phospholipid binding site. The purpose of this study was to investigate directly the role of the C2 domain in phospholipid binding. Twenty-six overlapping peptides, which span the entire C2 domain of FVIII, were synthesized. The ability of these peptides to inhibit the binding of purified human FVIII to immobilized phosphatidylserine was evaluated in an enzyme-linked immunosorbent assay. Three overlapping synthetic FVIII peptides, 2303-2317, 2305-2332, and 2308-2322, inhibited FVIII binding to phosphatidylserine by greater than 90% when tested at a concentration of 100 mumols/L. A fourth partially overlapping peptide, 2318-2332, inhibited FVIII binding by 65%. These results suggest that the area described by these peptides, residues 2303 to 2332, may play an important role in the mediation of FVIII binding to phospholipid.
Foster
P A
PA
Division of Experimental Hemostasis, Roon Research Center for Arteriosclerosis and Thrombosis, Scripps Clinic and Research Foundation, La Jolla, CA 92037.
Fulcher
C A
CA
Houghten
R A
RA
Zimmerman
T S
TS
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis*; D003175:Complement C2 / Q000032:analysis* / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005169:Factor VIII / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D010455:Peptides / Q000032:analysis / Q000494:pharmacology*; D010718:Phosphatidylserines / Q000378:metabolism*; D010743:Phospholipids / Q000378:metabolism
D000596:Amino Acids; D003175:Complement C2; D010455:Peptides; D010718:Phosphatidylserines; D010743:Phospholipids; D005169:Factor VIII