Kinetics of the inhibition of thrombin by hirudin
Research article published in Biochemistry (1986)
Abstract
The dissociation constant for hirudin was determined by varying the concentration of hirudin in the presence of a fixed concentration of thrombin and tripeptidyl p-nitroanilide substrate. The estimate of the dissociation constant determined in this manner displayed a dependence on the concentration of substrate which suggested the existence of two binding sites at which the substrate was able to compete with hirudin. A high-affinity site could be correlated with the binding of the substrate at the active site, and the other site had an affinity for the substrate that was 2 orders of magnitude lower. Extrapolation to zero substrate concentration yielded a value of 20 fM for the dissociation constant of hirudin at an ionic strength of 0.125. The dissociation constant for hirudin was markedly dependent on the ionic strength of the assay; it increased 20-fold when the ionic strength was increased from 0.1 to 0.4. This increase in dissociation constant was accompanied by a decrease in the rate with which hirudin associated with thrombin. This rate could be measured with a conventional recording spectrophotometer at higher ionic strength and was found to be independent of the binding of substrate at the active site.
Abstract sourced from PubMed (NCBI) for the cited record. See the original publication for the authoritative version.
Resumen
The dissociation constant for hirudin was determined by varying the concentration of hirudin in the presence of a fixed concentration of thrombin and tripeptidyl p-nitroanilide substrate.
Por qué esto importa para la hirudoterapia
Este estudio cuantificó la cinética de unión entre la hirudina y la trombina, informando una constante de disociación de 20 fM a una fuerza iónica de 0,125 y mostrando que esta constante aumentó 20 veces cuando la fuerza iónica se elevó de 0,1 a 0,4, acompañada por una disminución de la velocidad de asociación. Los resultados también sugirieron dos sitios de unión competitivos con el sustrato en la trombina. Debido a que la hirudina es un potente inhibidor de la trombina de relevancia directa para los mecanismos anticoagulantes subyacentes a la hirudoterapia, estos parámetros cinéticos precisos aportan detalles bioquímicos fundamentales sobre esa interacción molecular. Sin embargo, este trabajo se limita a mediciones de cinética de unión in vitro con material purificado realizadas con un espectrofotómetro; no contiene contexto fisiológico ni clínico, no involucra sanguijuelas ni secreciones salivales, y su conexión con la práctica terapéutica es indirecta.
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Añadido a la biblioteca ASH: March 18, 2026 · Última actualización del sitio: June 18, 2026