Selective inhibition of factor Xa in the prothrombinase complex by the carboxyl-terminal domain of antistasin.
Studies of antistasin, a potent factor Xa inhibitor with anticoagulant properties, were performed wherein the properties of the full-length antistasin polypeptide (ATS-119) were compared with the properties of forms of antistasin truncated at residue 116 (ATS-116) and residue 112 (ATS-112). ATS-119 was 40-fold more potent than ATS-112 in prolonging the activated partial thromboplastin time (APTT), whereas ATS-119 inhibited factor Xa 2.2-fold less avidly and about 5-fold more slowly than did ATS-112. The decreased reactivity of ATS-119 suggests that the carboxyl-terminal domain of ATS-119 stabilizes an ATS conformation with a reduced reactivity toward factor Xa. The observation that calcium ion increases the reactivity of ATS-119 but not that of ATS-112 suggests that calcium ion may disrupt interactions involving the carboxyl terminus of ATS-119. Interestingly, ATS-119 inhibited factor Xa in the prothrombinase complex 2-6-fold more potently and 2-3-fold faster than ATS-112. These differences in affinity and reactivity might well account for the greater effectiveness of ATS-119 in prolonging the APTT and suggest that the carboxyl-terminal domain of ATS-119 disrupts interactions involving phospholipid, factor Va, and prothrombin in the prothrombinase complex. The peptide RPKRKLIPRLS, corresponding to the carboxyl domain of ATS-119 prolonged the APTT and inhibited prothrombinase-catalyzed processing of prothrombin, but it failed to inhibit the catalytic activity of isolated factor Xa. Thus, this novel inhibitor appears to exert its inhibitory effects at a site removed from the active site of factor Xa.
Mao
S S
SS
Department of Biological Chemistry, Merck Research Laboratories, West Point, Pennsylvania 19486, USA.
Przysiecki
C T
CT
Krueger
J A
JA
Cooper
C M
CM
Lewis
S D
SD
Joyce
J
J
Lellis
C
C
Garsky
V M
VM
Sardana
M
M
Shafer
J A
JA
D000595:Amino Acid Sequence; D000818:Animals; D000925:Anticoagulants / Q000737:chemistry / Q000494:pharmacology*; D019556:COS Cells; D002384:Catalysis; D002417:Cattle; D065427:Factor Xa Inhibitors; D006801:Humans; D007447:Invertebrate Hormones / Q000737:chemistry / Q000494:pharmacology*; D007700:Kinetics; D008969:Molecular Sequence Data; D010314:Partial Thromboplastin Time; D010446:Peptide Fragments / Q000737:chemistry / Q000494:pharmacology*; D011516:Prothrombin / Q000378:metabolism; D018411:Spodoptera; D013925:Thromboplastin / Q000378:metabolism*
D000925:Anticoagulants; D065427:Factor Xa Inhibitors; D007447:Invertebrate Hormones; D010446:Peptide Fragments; C053155:antistasin; D011516:Prothrombin; D013925:Thromboplastin