Hirudin Fragments and Analogs
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Introduction
Hirudin is a kind of polypeptide with 65 or 66 amino acids, originally isolated from the medicinal leech Hirudo medicinalis. However, the recombinant forms of the inhibitor are now available. It can be a tight-binding and slow inhibitor of thrombin. Study has shown that the binding of a C-terminal fragment of hirudin induces a conformational change in thrombin. Thus, the hirudin fragments and analogs have been used to investigate the regions of thrombin and hirudin that interact with each other. The structure of hirudin fragments and analogs may provide a basis for the design of specific thrombin inhibitors.
Mechanism of action
The synthetic hirudin fragments from Asp (55) to Gln (65) (H-Asn-Asp-Gly-Asp (55)-Phe-Glu-Glu-Ile-Pro-Glu-Glu-Tyr (63)-Leu-Gln-COOH and acetyl-Asp (55)-Phe-Glu-Glu-Ile-Pro-Glu-Glu-Tyr (63)-Leu-Gln-COOH) are important in the interaction with thrombin. The thrombin generation depends on the availability of the prothrobinase complex. The complex consists of phospholipid and Ca2+. Although the factor V is activated by the factor Xa, it is much more rapidly activated by thrombin. This not only occurs with α-thrombin but also can occur with nonclotting γ-thrombin. While the hirudin completely blochs both clotting activity and factor V activation the dodecapeptide ‘hirugen’ blocks clotting but not factor V activation. Thus, clotting or thrombosis can conceivably be inhibited without completely preventing factor V activation, which would increase the risk of bleeding. The apolar disulfide knot of hirudin masks the catalytic site of thrombin by interacting with apolar regions surrounding the catalytic site. The anionic tail of hirudin interacts ionically with the anion-binding exosite of α-thrombin and combination of these two major interactions explains the high affinity binding of the α-thrombin-hirudin complex. Rather than the inhibition of clotting activity, blocking factor V activation in the amplification of prothrombinase formation explains the physiologic inhibition of thrombin generation caused by hirudun.
Application of Hirudin Fragments and Analogs
Hirudin has a blood anticoagulant property. This is fundamental for the leeches’ alimentary habit of hematophagy, since it keeps the blood flowing after the initial phlebotomy performed by the worm on the host’s skin. An interaction like that between hirudin and thrombin, which involves a number of salt bridges, will in general be sensitive to ionic strength. The hirudin-thrombin binding energy is sensitive to ionic strength, and the sensitivity is affected by the muta- tions in the acidic C-terminal region of hirudin. However, the presence of sodium and chloride binding sites on thrombin complicates the interpretation of these salt sensitivity measurements when the buffer contains sodium.
References
- Li, J., Liu, F., Qin, Y., He, J., Xiong, Z., Deng, G., & Li, Q. (2017). A novel natural hirudin facilitated anti-clotting polylactide membrane via hydrogen bonding interaction. Journal of Membrane Science, 523, 505-514.
- Sharp, K. A. (1996). Electrostatic interactions in hirudin-thrombin binding. Biophysical chemistry, 61(1), 37-49.
- Fenton II, J. W., Villanueva, G. B., Ofosu, F. A., & Maraganore, J. M. (1991). Thrombin inhibition by hirudin: how hirudin inhibits thrombin. Pathophysiology of Haemostasis and Thrombosis, 21(Suppl. 1), 27-31.
