Bordonein-L, a new L-amino acid oxidase from Crotalus durissus terrificus snake venom: isolation, preliminary characterization and enzyme stability
BackgroundCrotalus durissus terrificus venom (CdtV) is one of the most studied snake venoms in Brazil. Despite presenting several well known proteins, its L-amino acid oxidase (LAAO) has not been studied previously. This study aimed to isolate, characterize and evaluate the enzyme stability of bordo...
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ftdoajarticles:oai:doaj.org/article:7651625ddd804570822ef779de96bb41 2023-05-15T15:14:32+02:00 Bordonein-L, a new L-amino acid oxidase from Crotalus durissus terrificus snake venom: isolation, preliminary characterization and enzyme stability Karla C. F. Bordon Gisele A. Wiezel Hamilton Cabral Eliane C. Arantes 2015-09-01T00:00:00Z https://doi.org/10.1186/s40409-015-0025-8 https://doaj.org/article/7651625ddd804570822ef779de96bb41 EN eng SciELO http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992015000100335&lng=en&tlng=en https://doaj.org/toc/1678-9199 1678-9199 doi:10.1186/s40409-015-0025-8 https://doaj.org/article/7651625ddd804570822ef779de96bb41 Journal of Venomous Animals and Toxins including Tropical Diseases, Vol 21, Iss 0 (2015) Crotalus durissus terrificus L-amino acid oxidase Rattlesnake Enzyme activity Enzyme stability Chromatography Snake venom Yellow venom Stabilization Arctic medicine. Tropical medicine RC955-962 Toxicology. Poisons RA1190-1270 Zoology QL1-991 article 2015 ftdoajarticles https://doi.org/10.1186/s40409-015-0025-8 2022-12-31T01:07:41Z BackgroundCrotalus durissus terrificus venom (CdtV) is one of the most studied snake venoms in Brazil. Despite presenting several well known proteins, its L-amino acid oxidase (LAAO) has not been studied previously. This study aimed to isolate, characterize and evaluate the enzyme stability of bordonein-L, an LAAO from CdtV.Methods The enzyme was isolated through cation exchange, gel filtration and affinity chromatography, followed by a reversed-phase fast protein liquid chromatography to confirm its purity. Subsequently, its N-terminal amino acid sequence was determined by Edman degradation. The enzyme activity and stability were evaluated by a microplate colorimetric assay and the molecular mass was estimated by SDS-PAGE using periodic acid-Schiff staining and determined by mass spectrometry.Results The first 39 N-terminal amino acid residues exhibited high identity with other snake venom L-amino acid oxidases. Bordonein-L is a homodimer glycoprotein of approximately 101 kDa evaluated by gel filtration. Its monomer presents around 53 kDa estimated by SDS-PAGE and 58,702 Da determined by MALDI-TOF mass spectrometry. The enzyme exhibited maximum activity at pH 7.0 and lost about 50 % of its activity after five days of storage at 4 °C. Bordonein-L’s activity was higher than the control when stored in 2.8 % mannitol or 8.5 % sucrose.Conclusions This research is pioneering in its isolation, characterization and enzyme stability evaluation of an LAAO from CdtV, denominated bordonein-L. These results are important because they increase the knowledge about stabilization of LAAOs, aiming to increase their shelf life. Since the maintenance of enzymatic activity after long periods of storage is essential to enable their biotechnological use as well as their functional studies. Article in Journal/Newspaper Arctic Directory of Open Access Journals: DOAJ Articles Arctic Journal of Venomous Animals and Toxins including Tropical Diseases 21 1 |
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Directory of Open Access Journals: DOAJ Articles |
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ftdoajarticles |
language |
English |
topic |
Crotalus durissus terrificus L-amino acid oxidase Rattlesnake Enzyme activity Enzyme stability Chromatography Snake venom Yellow venom Stabilization Arctic medicine. Tropical medicine RC955-962 Toxicology. Poisons RA1190-1270 Zoology QL1-991 |
spellingShingle |
Crotalus durissus terrificus L-amino acid oxidase Rattlesnake Enzyme activity Enzyme stability Chromatography Snake venom Yellow venom Stabilization Arctic medicine. Tropical medicine RC955-962 Toxicology. Poisons RA1190-1270 Zoology QL1-991 Karla C. F. Bordon Gisele A. Wiezel Hamilton Cabral Eliane C. Arantes Bordonein-L, a new L-amino acid oxidase from Crotalus durissus terrificus snake venom: isolation, preliminary characterization and enzyme stability |
topic_facet |
Crotalus durissus terrificus L-amino acid oxidase Rattlesnake Enzyme activity Enzyme stability Chromatography Snake venom Yellow venom Stabilization Arctic medicine. Tropical medicine RC955-962 Toxicology. Poisons RA1190-1270 Zoology QL1-991 |
description |
BackgroundCrotalus durissus terrificus venom (CdtV) is one of the most studied snake venoms in Brazil. Despite presenting several well known proteins, its L-amino acid oxidase (LAAO) has not been studied previously. This study aimed to isolate, characterize and evaluate the enzyme stability of bordonein-L, an LAAO from CdtV.Methods The enzyme was isolated through cation exchange, gel filtration and affinity chromatography, followed by a reversed-phase fast protein liquid chromatography to confirm its purity. Subsequently, its N-terminal amino acid sequence was determined by Edman degradation. The enzyme activity and stability were evaluated by a microplate colorimetric assay and the molecular mass was estimated by SDS-PAGE using periodic acid-Schiff staining and determined by mass spectrometry.Results The first 39 N-terminal amino acid residues exhibited high identity with other snake venom L-amino acid oxidases. Bordonein-L is a homodimer glycoprotein of approximately 101 kDa evaluated by gel filtration. Its monomer presents around 53 kDa estimated by SDS-PAGE and 58,702 Da determined by MALDI-TOF mass spectrometry. The enzyme exhibited maximum activity at pH 7.0 and lost about 50 % of its activity after five days of storage at 4 °C. Bordonein-L’s activity was higher than the control when stored in 2.8 % mannitol or 8.5 % sucrose.Conclusions This research is pioneering in its isolation, characterization and enzyme stability evaluation of an LAAO from CdtV, denominated bordonein-L. These results are important because they increase the knowledge about stabilization of LAAOs, aiming to increase their shelf life. Since the maintenance of enzymatic activity after long periods of storage is essential to enable their biotechnological use as well as their functional studies. |
format |
Article in Journal/Newspaper |
author |
Karla C. F. Bordon Gisele A. Wiezel Hamilton Cabral Eliane C. Arantes |
author_facet |
Karla C. F. Bordon Gisele A. Wiezel Hamilton Cabral Eliane C. Arantes |
author_sort |
Karla C. F. Bordon |
title |
Bordonein-L, a new L-amino acid oxidase from Crotalus durissus terrificus snake venom: isolation, preliminary characterization and enzyme stability |
title_short |
Bordonein-L, a new L-amino acid oxidase from Crotalus durissus terrificus snake venom: isolation, preliminary characterization and enzyme stability |
title_full |
Bordonein-L, a new L-amino acid oxidase from Crotalus durissus terrificus snake venom: isolation, preliminary characterization and enzyme stability |
title_fullStr |
Bordonein-L, a new L-amino acid oxidase from Crotalus durissus terrificus snake venom: isolation, preliminary characterization and enzyme stability |
title_full_unstemmed |
Bordonein-L, a new L-amino acid oxidase from Crotalus durissus terrificus snake venom: isolation, preliminary characterization and enzyme stability |
title_sort |
bordonein-l, a new l-amino acid oxidase from crotalus durissus terrificus snake venom: isolation, preliminary characterization and enzyme stability |
publisher |
SciELO |
publishDate |
2015 |
url |
https://doi.org/10.1186/s40409-015-0025-8 https://doaj.org/article/7651625ddd804570822ef779de96bb41 |
geographic |
Arctic |
geographic_facet |
Arctic |
genre |
Arctic |
genre_facet |
Arctic |
op_source |
Journal of Venomous Animals and Toxins including Tropical Diseases, Vol 21, Iss 0 (2015) |
op_relation |
http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992015000100335&lng=en&tlng=en https://doaj.org/toc/1678-9199 1678-9199 doi:10.1186/s40409-015-0025-8 https://doaj.org/article/7651625ddd804570822ef779de96bb41 |
op_doi |
https://doi.org/10.1186/s40409-015-0025-8 |
container_title |
Journal of Venomous Animals and Toxins including Tropical Diseases |
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21 |
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1 |
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1766344979857276928 |