Photoaffinity labeling of functionally different lysine-binding sites in human plasminogen and plasmin.
Photoaffinity labeling of human plasmin using 4-azidobenzoylglycyl-L-lysine inhibits clot lysis activity, while the activity toward the active-site titrant, p-nitrophenyl-p'-guanidinobenzoate, or alpha-casein are maintained. Photoaffinity labeling of native Glu-plasminogen with the same reagent causes incorporation of approximately 1.5 mol label per mol plasminogen. This labeled plasminogen can be activated to plasmin by either urokinase or streptokinase. The resulting plasmin has full clot lysis activity and can be subsequently photoaffinity labeled with a loss of clot lysis activity. The rate of activation of labeled plasminogen by urokinase is increased relative to that of native plasminogen. epsilon-Aminocaproic acid blocks incorporation of photoaffinity label into both plasminogen and plasmin, indicating that the labeling is specific to the lysine-binding sites. The labels are located in the kringle 1+2+3 fragment in either photoaffinity-labeled plasminogen or plasmin. These results indicate that the specific lysine-binding site blocked in plasmin acts in concert with the active-site in binding and using fibrin as a substrate. This clot lysis regulating site is not available for labeling in plasminogen, but is exposed or changed upon activation to plasmin. The different lysine-binding sites labeled in plasminogen may regulate the conformation of the molecule as evidence by an enhanced rate of activation to plasmin.
Ryan
T J
TJ
Keegan
M C
MC
D000345:Affinity Labels / Q000378:metabolism*; D000818:Animals; D001386:Azides / Q000378:metabolism; D001665:Binding Sites; D002417:Cattle; D004151:Dipeptides / Q000378:metabolism; D005341:Fibrinolysin / Q000378:metabolism*; D005342:Fibrinolysis / Q000187:drug effects; D006801:Humans; D008239:Lysine / Q000378:metabolism*; D010777:Photochemistry; D010958:Plasminogen / Q000378:metabolism*; D013300:Streptokinase / Q000378:metabolism; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism
D000345:Affinity Labels; D001386:Azides; D004151:Dipeptides; C046123:4-azidobenzoylglycyllysine; D010958:Plasminogen; D013300:Streptokinase; D005341:Fibrinolysin; D014568:Urokinase-Type Plasminogen Activator; D008239:Lysine