Urokinase binds to a plasminogen activator inhibitor type-2-like molecule in placental microvillous membranes.
Placental microvillous membranes exhibited saturable binding of urokinase-type plasminogen activator with plateau achieved by 30 min at 4 degrees C and 10 min at 37 degrees C. The binding was essentially irreversible. The capacity was about 8 pmol urokinase per mg membrane protein. Half-maximal displacement of 125I-labelled urokinase was achieved with about 1.0 nM unlabelled urokinase when using 75 micrograms membrane protein/ml. 125I-labelled urokinase did not bind when treated with diisopropylfluorophosphate to block the catalytic activity. Single-chain urokinase (prourokinase), devoid of catalytic activity, did not bind. Catalytically active tissue-type plasminogen activator did compete with 125I-labelled urokinase for binding although less efficiently than urokinase. Binding activity remained in the 100,000 x g pellet after treatment of the membranes with 3 M KCl, alkaline stripping at pH 12 or extraction by the detergent Triton X-100. The binding was essentially blocked by antibodies against plasminogen activator inhibitor-type-2 (PAI-2). Sodium dodecyl sulfate polyacrylamide gel electrophoresis of solubilized membranes with bound 125I-labelled urokinase showed that the urokinase-PAI-2 complexes largely migrated in fractions corresponding to a very large Mr although no clearly defined peaks were observed. It is suggested that PAI-2 occurs in a form anchored to syncytiotrophoblast microvilli, possibly to the cytoskeleton.
Jensen
P H
PH
Institute of Physiology, University of Aarhus, Denmark.
Nykjaer
A
A
Andreasen
P A
PA
Lund
L R
LR
Astedt
B
B
Lecander
I
I
Gliemann
J
J
D002824:Chorionic Villi / Q000378:metabolism; D005260:Female; D006801:Humans; D007700:Kinetics; D008871:Microvilli / Q000378:metabolism*; D010920:Placenta / Q000378:metabolism*; D015849:Plasminogen Inactivators / Q000378:metabolism*; D011247:Pregnancy; D011485:Protein Binding; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism*
D015849:Plasminogen Inactivators; D014568:Urokinase-Type Plasminogen Activator