Beta-hydroxyaspartic acid in the first epidermal growth factor-like domain of protein C. Its role in Ca2+ binding and biological activity.
Protein C is a vitamin K-dependent regulator of blood coagulation. It has beta-hydroxyaspartic acid in position 71 which is in the first of its two domains that are homologous to epidermal growth factor (EGF). This region has recently been demonstrated to have a Ca2+ binding site with a Kd of approximately 100 microM. Recombinant human protein C, expressed in mammalian tissue culture, had full biological activity and contained beta-hydroxyaspartic acid. Furthermore, it had a Ca2+-dependent epitope in the EGF-like domain, recognized by a monoclonal antibody. In contrast, a mutant recombinant human protein C in which beta-hydroxyaspartic acid had been replaced with glutamic acid in position 71 did not have the Ca2+-dependent epitope, and its biological activity was reduced to about 10% of normal. Fab' fragments of this antibody inhibited the anticoagulant activity of plasma-derived activated protein C, apparently by interfering with the interaction between activated protein C and its cofactor, protein S. The latter contains four tandemly arranged EGF homology domains. We propose that beta-hydroxyaspartic acid is directly involved in Ca2+ binding in protein C and in related proteins and that protein C interacts with protein S by means of its EGF homology regions.
Ohlin
A K
AK
Department of Clinical Chemistry, University of Lund, Malmö General Hospital, Sweden.
Landes
G
G
Bourdon
P
P
Oppenheimer
C
C
Wydro
R
R
Stenflo
J
J
D000911:Antibodies, Monoclonal; D001224:Aspartic Acid / Q000031:analogs & derivatives / Q000032:analysis; D001483:Base Sequence; D002118:Calcium / Q000378:metabolism; D004815:Epidermal Growth Factor / Q000032:analysis*; D000939:Epitopes / Q000032:analysis; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D011486:Protein C / Q000032:analysis*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D011486:Protein C; C011274:3-hydroxyaspartic acid; D001224:Aspartic Acid; D004815:Epidermal Growth Factor; D002118:Calcium