Structure and specificity studies on batroxobin, a snake venom derived thrombin-like enzyme
Structure and specificity studies on batroxobin, a snake venom derived thrombin-like enzyme
Batroxobin derived from two sources, Bothrops atrox and Bothrops moojeni, two subspecies of the same Bothrops Atrox species, were characterised. It was not possible to identify any difference between them in terms of mass and N-terminal sequence analysis. However, on reaction with various fibrinogens it was evident that the batroxobin derived from Bothrops atrox was twice as active as the batroxobin derived from Bothrops moojeni. Both batroxobins had a preference for fibrinogens derived from rat and mouse plasma, the natural prey of Bothrops Atrox pit vipers.
Sequence analysis of the fibrinogens indicated some differences in the sequences around the scissile bond of fibrinopeptide A, which could account for differences observed in specificity between the batroxobins and thrombin. It was found that Batroxobin has an absolute requirement for a proline residue at P2' position, unlike thrombin which can tolerate histidine. It was also evident that batroxobin has a preference to an aspartic acid at the P3 position, whereas thrombin prefers a glycine in this position. The synthesis of a novel inhibitor with a reduced peptide bond showed that both batroxobin and thrombin have a requirement for a carbonyl group at the scissile bond not only for cleavage, but also for recognition.
In an attempt to find a structural explanation for the observations, homology modelling was performed. A model produced from the structures of porcine glandular kallikrein and TSV-PA was finally decided upon. The sequences of various peptides were modelled into the active site cleft of the batroxobin model. This work showed that the preference of batroxobin for an aspartic acid at P3 could be due to its ability to form a stabilising interaction with a lysine in the active site cleft, which is not present in thrombin. It also indicated that batroxobin cannot tolerate histidine at the P2' position due to the highly positive nature of the enzyme at that point.
University of Southampton
1999
Earps, Lorraine
(1999)
Structure and specificity studies on batroxobin, a snake venom derived thrombin-like enzyme.
University of Southampton, Doctoral Thesis.
Record type:
Thesis
(Doctoral)
Abstract
Batroxobin derived from two sources, Bothrops atrox and Bothrops moojeni, two subspecies of the same Bothrops Atrox species, were characterised. It was not possible to identify any difference between them in terms of mass and N-terminal sequence analysis. However, on reaction with various fibrinogens it was evident that the batroxobin derived from Bothrops atrox was twice as active as the batroxobin derived from Bothrops moojeni. Both batroxobins had a preference for fibrinogens derived from rat and mouse plasma, the natural prey of Bothrops Atrox pit vipers.
Sequence analysis of the fibrinogens indicated some differences in the sequences around the scissile bond of fibrinopeptide A, which could account for differences observed in specificity between the batroxobins and thrombin. It was found that Batroxobin has an absolute requirement for a proline residue at P2' position, unlike thrombin which can tolerate histidine. It was also evident that batroxobin has a preference to an aspartic acid at the P3 position, whereas thrombin prefers a glycine in this position. The synthesis of a novel inhibitor with a reduced peptide bond showed that both batroxobin and thrombin have a requirement for a carbonyl group at the scissile bond not only for cleavage, but also for recognition.
In an attempt to find a structural explanation for the observations, homology modelling was performed. A model produced from the structures of porcine glandular kallikrein and TSV-PA was finally decided upon. The sequences of various peptides were modelled into the active site cleft of the batroxobin model. This work showed that the preference of batroxobin for an aspartic acid at P3 could be due to its ability to form a stabilising interaction with a lysine in the active site cleft, which is not present in thrombin. It also indicated that batroxobin cannot tolerate histidine at the P2' position due to the highly positive nature of the enzyme at that point.
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Published date: 1999
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Local EPrints ID: 463975
URI: http://eprints.soton.ac.uk/id/eprint/463975
PURE UUID: 9f5832ed-cffa-43b6-8be7-3cc3b0a4a444
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Date deposited: 04 Jul 2022 20:59
Last modified: 04 Jul 2022 20:59
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Author:
Lorraine Earps
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