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Identification and Mechanistic Analysis of a Novel Tick-Derived Inhibitor of Thrombin.

Jablonka W, Kotsyfakis M, Mizurini DM, Monteiro RQ, Lukszo J, Drake SK, Ribeiro JM, Andersen JF - PLoS ONE (2015)

Bottom Line: Hyalomin-1 is cleaved at a canonical thrombin cleavage site but the cleaved products do not inhibit coagulation.A peptide combining the N-terminal parts of the molecule with the cleavage region did not interact strongly with thrombin, but a 24-residue fragment containing the cleavage region and the C-terminal fragment inhibited the enzyme in a competitive manner and also inhibited coagulation of plasma.These results suggest that the peptide acts by binding to the active site as well as exosite I or the autolysis loop of thrombin.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Malaria and Vector Research, NIAID, National Institutes of Health, Rockville, Maryland, United States of America.

ABSTRACT
A group of peptides from the salivary gland of the tick Hyalomma marginatum rufipes, a vector of Crimean Congo hemorrhagic fever show weak similarity to the madanins, a group of thrombin-inhibitory peptides from a second tick species, Haemaphysalis longicornis. We have evaluated the anti-serine protease activity of one of these H. marginatum peptides that has been given the name hyalomin-1. Hyalomin-1 was found to be a selective inhibitor of thrombin, blocking coagulation of plasma and inhibiting S2238 hydrolysis in a competitive manner with an inhibition constant (Ki) of 12 nM at an ionic strength of 150 mM. It also blocks the thrombin-mediated activation of coagulation factor XI, thrombin-mediated platelet aggregation, and the activation of coagulation factor V by thrombin. Hyalomin-1 is cleaved at a canonical thrombin cleavage site but the cleaved products do not inhibit coagulation. However, the C-terminal cleavage product showed non-competitive inhibition of S2238 hydrolysis. A peptide combining the N-terminal parts of the molecule with the cleavage region did not interact strongly with thrombin, but a 24-residue fragment containing the cleavage region and the C-terminal fragment inhibited the enzyme in a competitive manner and also inhibited coagulation of plasma. These results suggest that the peptide acts by binding to the active site as well as exosite I or the autolysis loop of thrombin. Injection of 2.5 mg/kg of hyalomin-1 increased arterial occlusion time in a mouse model of thrombosis, suggesting this peptide could be a candidate for clinical use as an antithrombotic.

No MeSH data available.


Related in: MedlinePlus

Hyalomin-1 exhibits antithrombotic activity in vivo.(A) Occlusion of the carotid artery in BALB/c mice induced by FeCl3 and measured by laser Doppler flowmetry after injection of vehicle alone or hyalomin-1 at doses of 1 and 2.5 mg/kg. (B) Bleeding from the tail of BALB/c mice after injection of vehicle alone or vehicle plus hyalomin-1 at a dose of 2.5mg/kg. Statistical significance levels: * P < 0.05, ** P < 0.001.
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pone.0133991.g007: Hyalomin-1 exhibits antithrombotic activity in vivo.(A) Occlusion of the carotid artery in BALB/c mice induced by FeCl3 and measured by laser Doppler flowmetry after injection of vehicle alone or hyalomin-1 at doses of 1 and 2.5 mg/kg. (B) Bleeding from the tail of BALB/c mice after injection of vehicle alone or vehicle plus hyalomin-1 at a dose of 2.5mg/kg. Statistical significance levels: * P < 0.05, ** P < 0.001.

Mentions: In order to assess the potential of hyalomin-1 as an antithrombotic agent, in vivo thrombosis assays were carried out in mice. Injury of the carotid artery was induced by ferric chloride after intravenous injection of hyalomin-1 at concentrations of 1 or 2.5 mg/kg and the time to arterial occlusion was measured. Hyalomin-1 produced a detectable delay in occlusion at 1 mg/kg and a statistically significant 1.6 fold delay at 2.5 mg/kg peptide (Fig 7A).


Identification and Mechanistic Analysis of a Novel Tick-Derived Inhibitor of Thrombin.

Jablonka W, Kotsyfakis M, Mizurini DM, Monteiro RQ, Lukszo J, Drake SK, Ribeiro JM, Andersen JF - PLoS ONE (2015)

Hyalomin-1 exhibits antithrombotic activity in vivo.(A) Occlusion of the carotid artery in BALB/c mice induced by FeCl3 and measured by laser Doppler flowmetry after injection of vehicle alone or hyalomin-1 at doses of 1 and 2.5 mg/kg. (B) Bleeding from the tail of BALB/c mice after injection of vehicle alone or vehicle plus hyalomin-1 at a dose of 2.5mg/kg. Statistical significance levels: * P < 0.05, ** P < 0.001.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4526366&req=5

pone.0133991.g007: Hyalomin-1 exhibits antithrombotic activity in vivo.(A) Occlusion of the carotid artery in BALB/c mice induced by FeCl3 and measured by laser Doppler flowmetry after injection of vehicle alone or hyalomin-1 at doses of 1 and 2.5 mg/kg. (B) Bleeding from the tail of BALB/c mice after injection of vehicle alone or vehicle plus hyalomin-1 at a dose of 2.5mg/kg. Statistical significance levels: * P < 0.05, ** P < 0.001.
Mentions: In order to assess the potential of hyalomin-1 as an antithrombotic agent, in vivo thrombosis assays were carried out in mice. Injury of the carotid artery was induced by ferric chloride after intravenous injection of hyalomin-1 at concentrations of 1 or 2.5 mg/kg and the time to arterial occlusion was measured. Hyalomin-1 produced a detectable delay in occlusion at 1 mg/kg and a statistically significant 1.6 fold delay at 2.5 mg/kg peptide (Fig 7A).

Bottom Line: Hyalomin-1 is cleaved at a canonical thrombin cleavage site but the cleaved products do not inhibit coagulation.A peptide combining the N-terminal parts of the molecule with the cleavage region did not interact strongly with thrombin, but a 24-residue fragment containing the cleavage region and the C-terminal fragment inhibited the enzyme in a competitive manner and also inhibited coagulation of plasma.These results suggest that the peptide acts by binding to the active site as well as exosite I or the autolysis loop of thrombin.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Malaria and Vector Research, NIAID, National Institutes of Health, Rockville, Maryland, United States of America.

ABSTRACT
A group of peptides from the salivary gland of the tick Hyalomma marginatum rufipes, a vector of Crimean Congo hemorrhagic fever show weak similarity to the madanins, a group of thrombin-inhibitory peptides from a second tick species, Haemaphysalis longicornis. We have evaluated the anti-serine protease activity of one of these H. marginatum peptides that has been given the name hyalomin-1. Hyalomin-1 was found to be a selective inhibitor of thrombin, blocking coagulation of plasma and inhibiting S2238 hydrolysis in a competitive manner with an inhibition constant (Ki) of 12 nM at an ionic strength of 150 mM. It also blocks the thrombin-mediated activation of coagulation factor XI, thrombin-mediated platelet aggregation, and the activation of coagulation factor V by thrombin. Hyalomin-1 is cleaved at a canonical thrombin cleavage site but the cleaved products do not inhibit coagulation. However, the C-terminal cleavage product showed non-competitive inhibition of S2238 hydrolysis. A peptide combining the N-terminal parts of the molecule with the cleavage region did not interact strongly with thrombin, but a 24-residue fragment containing the cleavage region and the C-terminal fragment inhibited the enzyme in a competitive manner and also inhibited coagulation of plasma. These results suggest that the peptide acts by binding to the active site as well as exosite I or the autolysis loop of thrombin. Injection of 2.5 mg/kg of hyalomin-1 increased arterial occlusion time in a mouse model of thrombosis, suggesting this peptide could be a candidate for clinical use as an antithrombotic.

No MeSH data available.


Related in: MedlinePlus