Characterization of rat factors X and Xa: demonstration of factor Xa in rat plasma.
We have found that rat plasma corrected the non-activated PT of human normal or factor-X deficient plasma, and the factor Xa-like activity being constantly detected in every 1 ml of blood collected via the cannulated carotid artery of rats. The present study was undertaken to characterize the factor Xa-like activity in rat plasma by preparing rat factor X and a monoclonal antibody against it. Factor X was purified from a BaCl2 eluate of rat plasma by chromatographies on columns of DEAE-Sepharose CL-6B and Sulfate Cellulofine or on a column of Affi-Gel 10 conjugated with a monoclonal antibody against rat factor X. Factor Xa-like activity in rat plasma was eliminated by the treatment of rat plasma with a monoclonal antibody which recognized the heavy chain portions of rat factors X and Xa. A kinetical study demonstrated that rat factor Xa was strongly inhibited by rat antithrombin III, with a Ki of 2.2 x 10(-11) M, in the presence of heparin. However, in the absence of heparin, the second order rate constant for the inhibition of rat factor Xa by rat antithrombin III was 2.6 x 10(4) M-1.min-1, which was one forty-third that for the inhibition of human factor Xa by human antithrombin III. Furthermore, rat factor Xa was resistant to the inhibition by rat alpha-1-antitrypsin and alpha-2-macroglobulin.(ABSTRACT TRUNCATED AT 250 WORDS)
Enjyoji
K
K
National Cardiovascular Center Research Institute, Osaka University.
Miyazaki
K
K
Kato
H
H
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D000990:Antithrombin III / Q000302:isolation & purification; D002417:Cattle; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005170:Factor X / Q000032:analysis*; D015951:Factor Xa / Q000032:analysis*; D005453:Fluorescence; D006801:Humans; D015151:Immunoblotting; D008969:Molecular Sequence Data; D011517:Prothrombin Time; D051381:Rats; D000515:alpha 1-Antitrypsin / Q000302:isolation & purification; D000511:alpha-Macroglobulins / Q000302:isolation & purification
D000911:Antibodies, Monoclonal; D000515:alpha 1-Antitrypsin; D000511:alpha-Macroglobulins; D000990:Antithrombin III; D005170:Factor X; D015951:Factor Xa