Antistasin, a leech-derived inhibitor of factor Xa. Kinetic analysis of enzyme inhibition and identification of the reactive site
Research article published in J Biol Chem (1989)
Abstract
Antistasin is a 119-amino acid protein isolated from the salivary glands of the Mexican leech Haementeria officinalis. The determination of the primary structure of antistasin revealed that the protein is highly disulfide-bonded with a 2-fold internal homology. Antistasin exhibits a potent anticoagulant activity purportedly due to the selective inhibition of Factor Xa (Tuszinsky, G. P., Gasic, T. B., and Gasic, G. J. (1987) J. Biol. chem. 262, 9718-9723). In the present study a detailed kinetic analysis of the inhibitory interaction between antistasin and Factor Xa was performed. In addition, the specificity of antistasin was examined by testing its ability to inhibit a variety of serine proteinases. Utilizing purified antistasin and a tripetidyl p-nitroanilide substrate, antistasin was shown to act as a reversible inhibitor of Factor Xa which exhibits slow-tight binding kinetics. Antistasin reacts stoichiometrically with Factor Xa with inhibition displaying a mixed, primarily competitive type. The inhibition is partial in the presence of Ca2+ and becomes complete in the absence of Ca2+. The estimated dissociation constant for the enzyme-inhibitor complex is between 0.31 and 0.62 nM. After binding to Factor Xa, antistasin is cleaved at a single site to yield a modified inhibitor. Automated gas-phase sequence analysis of the modified inhibitor indicates the arginine residue at position 34 in antistasin occupies the P1 position of the reactive site. These data indicate that the leech has evolved a highly selective and potent inhibitor of coagulation Factor Xa that shares several mechanistic similarities with other serine proteinase inhibitors.
Abstract sourced from PubMed (NCBI) for the cited record. See the original publication for the authoritative version.
Resumen
Antistasin is a 119-amino acid protein isolated from the salivary glands of the Mexican leech Haementeria officinalis.
Por qué esto importa para la hirudoterapia
Este estudio realizó un análisis cinético detallado de la antistasina, una proteína de 119 aminoácidos procedente de las glándulas salivales de la sanguijuela mexicana Haementeria officinalis, caracterizándola como un inhibidor potente, reversible y de unión lenta y firme del factor Xa, con una constante de disociación de 0,31–0,62 nM. Se identificó el sitio reactivo en la arginina-34, y la antistasina mostró selectividad entre las serina proteasas ensayadas, con una inhibición influenciada por la presencia de calcio. Esto es directamente relevante para el dominio de ASH, ya que caracteriza una proteína anticoagulante derivada de la secreción salival de la sanguijuela. ADVERTENCIA: se trata de un estudio bioquímico/enzimológico que utiliza proteínas purificadas in vitro; no se presentan datos in vivo, terapéuticos ni de hirudoterapia. Su valor para ASH reside en la ciencia molecular fundamental.
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Añadido a la biblioteca ASH: May 27, 2026 · Última actualización del sitio: 18 de junio de 2026