A Steady-state Template Model That Describes the Kinetics of Fibrin-stimulated [Glu1]- and [Lys78]Plasminogen Activation by Native tissue-type Plasminogen Activator and Variants That Lack Either the Finger or Kringle-2 Domain
| Autor: | Michael E. Nesheim, Anton J. G. Horrevoets, H. Pannekoek |
|---|---|
| Rok vydání: | 1997 |
| Předmět: |
Biochemistry
Tissue plasminogen activator Fibrin Enzyme activator Kringles medicine Humans Enzyme kinetics Molecular Biology Ternary complex biology Chemistry Activator (genetics) Plasminogen Zinc Fingers Templates Genetic Cell Biology Peptide Fragments Enzyme Activation Kinetics Models Chemical Tissue Plasminogen Activator biology.protein Steady state (chemistry) Plasminogen activator Algorithms medicine.drug |
| Zdroj: | Journal of Biological Chemistry. 272:2183-2191 |
| ISSN: | 0021-9258 |
| DOI: | 10.1074/jbc.272.4.2183 |
| Popis: | The kinetics of activation of both [Glu1]- and [Lys78]Plg(S741C-fluorescein by native (recombinant) tissue-type plasminogen activator and its deletion variants lacking either the finger or kringle-2 domain were measured by fluorescence within fully polymerized fibrin clots. The kinetics conform to the Michaelis-Menten equation at any fixed fibrin concentration so long as the plasminogen concentration is expressed as either the free or fibrin-bound, but not the total. The apparent kcat and Km values both vary systematically with the concentration of fibrin. Competition kinetics disclosed an active site-dependent interaction between t-Pa and [Glu1]Plg(S741C-fluorescein) in the presence, but not the absence, of fibrin. A steady-state template model having the rate equation v/[A]o = kcat(app).[Plg]/(Km(app) + [Plg]) was derived and used to interpret the data. The model indicates that catalytic efficiency is determined by the stability of the ternary activator-fibrin-plasminogen complex rather than the binding of the activator or plasminogen to fibrin. This implies that efforts to improve the enzymatic properties of t-PA might be more fruitfully directed at enhancing the stability of the ternary complex rather than fibrin binding. |
| Databáze: | OpenAIRE |
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