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Binding of tissue p...
Binding of tissue plasminogen activator to endothelial cells. The effect on functional properties. Localization of a ligand in the B-chain of tPA.
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Cheng, X F (author)
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- Brohlin, Maria, 1966- (author)
- Umeå universitet,Biomedicinsk laboratorievetenskap
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Pohl, G (author)
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Bäck, O (author)
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Wallén, P (author)
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(creator_code:org_t)
- 1995
- 1995
- English.
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In: Thrombosis Research. - 0049-3848 .- 1879-2472. ; 77:2, s. 149-64
- Related links:
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https://urn.kb.se/re...
Abstract
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- The binding of 125I-labelled tissue plasminogen activator (tPA), the tPA A- or B-chain to endothelial cells (EC) were studied in suspensions of cultured human umbilical vein EC (HUVEC) or immortalized microvascular EC (HMEC). By determinations of the concentration-dependent binding it was shown that both the A-chain and the B-chain, which were isolated after partial reduction of two-chain tPA, contain ligands for binding to EC. The affinity for the B-chain was much higher than for the A-chain according to Scatchard analysis (Kd 24 and 515 nM, respectively), whereas the number of binding sites was higher for the A-chain than for the B-chain (Bmax 8 x 10(5) and 1.2 x 10(5), respectively). There were no cross interactions between the A- and B-chains and their binding sites. The binding of tPA to EC induced an almost 100-fold increase of the activation rate when compared to the same amount of enzyme in free solution, which in contrast to the fibrin-induced stimulation was not inhibited by antibodies against fibrin. The enzymatic activity of the B-chain was much less affected by the association to the cells. Both tPA and the tPA B-chain were largely protected against inhibition by an excess plasminogen activator type-1 (PAI-1) when bound to EC, whereas the same amount of free tPA was totally inactivated. The competition studies strongly indicated that an N-terminal segment in the B-chain, AKHRRSPGER, may be the ligand part of the B-chain. It is interesting to note that this polypeptide segment also participates in a binding site for PAI-1, necessary for effective inhibition. This implies a possible competition between PAI-1 and a tPA-receptor for binding of tPA. High molecular weight urokinase had no quenching effect on the binding of the B-chain to EC.
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