Purification and characterization of the first gamma-phospholipase inhibitor (gamma PLI) from Bothrops jararaca snake serum
Autor(a) principal: | |
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Data de Publicação: | 2018 |
Outros Autores: | , , , , , , , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Repositório Institucional da UNESP |
Texto Completo: | http://dx.doi.org/10.1371/journal.pone.0193105 http://hdl.handle.net/11449/163937 |
Resumo: | Phospholipases A(2) (PLA(2)) are enzymes acting on the cell membrane phospholipids resulting in fatty acids and lysophospholipids and deconstructing the cell membrane. This protein is commonly found in snake venoms, causing tissue inflammation in the affected area. Evidence indicates that snakes have natural resistance to their own venom due to protective properties in plasma, that inhibit the action of proteins present in their venom. Given that, this study aimed to purify and characterize a gamma PLI from Bothrops jararaca serum, named gamma BjPLI. PLA(2) inhibitor was isolated using two chromatographic steps: an ion exchange column (DEAE), followed by an affinity column (crotoxin coupled to a CNBr-activated Sepharose resin). The purity and biochemical characterization of the isolated protein were analyzed by RP-HPLC, SEC, SDS-PAGE, circular dichroism and mass spectrometry. The ability to inhibit PLA(2) was determined by enzymatic activity, neutralization of paw edema and myonecrosis. The protein purity was confirmed by RP-HPLC and SEC, whilst an apparent molecular mass of 25 kDa and 20 kDa was obtained by SDS-PAGE, under reducing and non-reducing conditions, respectively. According to mass spectrometry analysis, this protein showed 72% and 68% of coverage when aligned to amino acid sequences of two proteins already described as PLIs. Thus, the inhibitory activity of enzymatic, edema and myonecrotic activities by gamma BjPLI suggests a role of this inhibitor for protection of these snakes against self-envenomation. |
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Purification and characterization of the first gamma-phospholipase inhibitor (gamma PLI) from Bothrops jararaca snake serumPhospholipases A(2) (PLA(2)) are enzymes acting on the cell membrane phospholipids resulting in fatty acids and lysophospholipids and deconstructing the cell membrane. This protein is commonly found in snake venoms, causing tissue inflammation in the affected area. Evidence indicates that snakes have natural resistance to their own venom due to protective properties in plasma, that inhibit the action of proteins present in their venom. Given that, this study aimed to purify and characterize a gamma PLI from Bothrops jararaca serum, named gamma BjPLI. PLA(2) inhibitor was isolated using two chromatographic steps: an ion exchange column (DEAE), followed by an affinity column (crotoxin coupled to a CNBr-activated Sepharose resin). The purity and biochemical characterization of the isolated protein were analyzed by RP-HPLC, SEC, SDS-PAGE, circular dichroism and mass spectrometry. The ability to inhibit PLA(2) was determined by enzymatic activity, neutralization of paw edema and myonecrosis. The protein purity was confirmed by RP-HPLC and SEC, whilst an apparent molecular mass of 25 kDa and 20 kDa was obtained by SDS-PAGE, under reducing and non-reducing conditions, respectively. According to mass spectrometry analysis, this protein showed 72% and 68% of coverage when aligned to amino acid sequences of two proteins already described as PLIs. Thus, the inhibitory activity of enzymatic, edema and myonecrotic activities by gamma BjPLI suggests a role of this inhibitor for protection of these snakes against self-envenomation.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Univ Sao Paulo, Inst Pesquisas Tecnol, Inst Butantan, Interunidades Biotecnol, Sao Paulo, SP, BrazilInst Butantan, Lab Herpetol, Sao Paulo, SP, BrazilUniv Estadual Paulista, Inst Biociencias Litoral Paulista, Sao Vicente, SP, BrazilUniv Fed Sao Paulo, Dept Bioquim, Sao Paulo, SP, BrazilUniv Estadual Paulista, Inst Biociencias Litoral Paulista, Sao Vicente, SP, BrazilFAPESP: 2012/19321-9FAPESP: 2016/03839-0FAPESP: 2013/05357-4FAPESP: 2014/11108-0FAPESP: 2017/01890-0FAPESP: 2017/16908-2FAPESP: 2013/12826-0Public Library ScienceUniversidade de São Paulo (USP)Inst ButantanUniversidade Estadual Paulista (Unesp)Universidade Federal de São Paulo (UNIFESP)Serino-Silva, CarolineMorais-Zani, KarenToyama, Marcos Hikari [UNESP]Toyama, Daniela de Oliveira [UNESP]Gaeta, Henrique Hesse [UNESP]Bittencourt Rodrigues, Caroline FabriAguiar, Weslei da SilvaTashima, Alexandre KeijiGrego, Kathleen FernandesTanaka-Azevedo, Anita Mitico2018-11-26T17:48:29Z2018-11-26T17:48:29Z2018-03-05info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/article16application/pdfhttp://dx.doi.org/10.1371/journal.pone.0193105Plos One. San Francisco: Public Library Science, v. 13, n. 3, 16 p., 2018.1932-6203http://hdl.handle.net/11449/16393710.1371/journal.pone.0193105WOS:000426548200004WOS000426548200004.pdf8573195327542061Web of Sciencereponame:Repositório Institucional da UNESPinstname:Universidade Estadual Paulista (UNESP)instacron:UNESPengPlos One1,164info:eu-repo/semantics/openAccess2023-10-13T06:04:07Zoai:repositorio.unesp.br:11449/163937Repositório InstitucionalPUBhttp://repositorio.unesp.br/oai/requestopendoar:29462024-08-05T14:44:22.790909Repositório Institucional da UNESP - Universidade Estadual Paulista (UNESP)false |
dc.title.none.fl_str_mv |
Purification and characterization of the first gamma-phospholipase inhibitor (gamma PLI) from Bothrops jararaca snake serum |
title |
Purification and characterization of the first gamma-phospholipase inhibitor (gamma PLI) from Bothrops jararaca snake serum |
spellingShingle |
Purification and characterization of the first gamma-phospholipase inhibitor (gamma PLI) from Bothrops jararaca snake serum Serino-Silva, Caroline |
title_short |
Purification and characterization of the first gamma-phospholipase inhibitor (gamma PLI) from Bothrops jararaca snake serum |
title_full |
Purification and characterization of the first gamma-phospholipase inhibitor (gamma PLI) from Bothrops jararaca snake serum |
title_fullStr |
Purification and characterization of the first gamma-phospholipase inhibitor (gamma PLI) from Bothrops jararaca snake serum |
title_full_unstemmed |
Purification and characterization of the first gamma-phospholipase inhibitor (gamma PLI) from Bothrops jararaca snake serum |
title_sort |
Purification and characterization of the first gamma-phospholipase inhibitor (gamma PLI) from Bothrops jararaca snake serum |
author |
Serino-Silva, Caroline |
author_facet |
Serino-Silva, Caroline Morais-Zani, Karen Toyama, Marcos Hikari [UNESP] Toyama, Daniela de Oliveira [UNESP] Gaeta, Henrique Hesse [UNESP] Bittencourt Rodrigues, Caroline Fabri Aguiar, Weslei da Silva Tashima, Alexandre Keiji Grego, Kathleen Fernandes Tanaka-Azevedo, Anita Mitico |
author_role |
author |
author2 |
Morais-Zani, Karen Toyama, Marcos Hikari [UNESP] Toyama, Daniela de Oliveira [UNESP] Gaeta, Henrique Hesse [UNESP] Bittencourt Rodrigues, Caroline Fabri Aguiar, Weslei da Silva Tashima, Alexandre Keiji Grego, Kathleen Fernandes Tanaka-Azevedo, Anita Mitico |
author2_role |
author author author author author author author author author |
dc.contributor.none.fl_str_mv |
Universidade de São Paulo (USP) Inst Butantan Universidade Estadual Paulista (Unesp) Universidade Federal de São Paulo (UNIFESP) |
dc.contributor.author.fl_str_mv |
Serino-Silva, Caroline Morais-Zani, Karen Toyama, Marcos Hikari [UNESP] Toyama, Daniela de Oliveira [UNESP] Gaeta, Henrique Hesse [UNESP] Bittencourt Rodrigues, Caroline Fabri Aguiar, Weslei da Silva Tashima, Alexandre Keiji Grego, Kathleen Fernandes Tanaka-Azevedo, Anita Mitico |
description |
Phospholipases A(2) (PLA(2)) are enzymes acting on the cell membrane phospholipids resulting in fatty acids and lysophospholipids and deconstructing the cell membrane. This protein is commonly found in snake venoms, causing tissue inflammation in the affected area. Evidence indicates that snakes have natural resistance to their own venom due to protective properties in plasma, that inhibit the action of proteins present in their venom. Given that, this study aimed to purify and characterize a gamma PLI from Bothrops jararaca serum, named gamma BjPLI. PLA(2) inhibitor was isolated using two chromatographic steps: an ion exchange column (DEAE), followed by an affinity column (crotoxin coupled to a CNBr-activated Sepharose resin). The purity and biochemical characterization of the isolated protein were analyzed by RP-HPLC, SEC, SDS-PAGE, circular dichroism and mass spectrometry. The ability to inhibit PLA(2) was determined by enzymatic activity, neutralization of paw edema and myonecrosis. The protein purity was confirmed by RP-HPLC and SEC, whilst an apparent molecular mass of 25 kDa and 20 kDa was obtained by SDS-PAGE, under reducing and non-reducing conditions, respectively. According to mass spectrometry analysis, this protein showed 72% and 68% of coverage when aligned to amino acid sequences of two proteins already described as PLIs. Thus, the inhibitory activity of enzymatic, edema and myonecrotic activities by gamma BjPLI suggests a role of this inhibitor for protection of these snakes against self-envenomation. |
publishDate |
2018 |
dc.date.none.fl_str_mv |
2018-11-26T17:48:29Z 2018-11-26T17:48:29Z 2018-03-05 |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/article |
format |
article |
status_str |
publishedVersion |
dc.identifier.uri.fl_str_mv |
http://dx.doi.org/10.1371/journal.pone.0193105 Plos One. San Francisco: Public Library Science, v. 13, n. 3, 16 p., 2018. 1932-6203 http://hdl.handle.net/11449/163937 10.1371/journal.pone.0193105 WOS:000426548200004 WOS000426548200004.pdf 8573195327542061 |
url |
http://dx.doi.org/10.1371/journal.pone.0193105 http://hdl.handle.net/11449/163937 |
identifier_str_mv |
Plos One. San Francisco: Public Library Science, v. 13, n. 3, 16 p., 2018. 1932-6203 10.1371/journal.pone.0193105 WOS:000426548200004 WOS000426548200004.pdf 8573195327542061 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
Plos One 1,164 |
dc.rights.driver.fl_str_mv |
info:eu-repo/semantics/openAccess |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
16 application/pdf |
dc.publisher.none.fl_str_mv |
Public Library Science |
publisher.none.fl_str_mv |
Public Library Science |
dc.source.none.fl_str_mv |
Web of Science reponame:Repositório Institucional da UNESP instname:Universidade Estadual Paulista (UNESP) instacron:UNESP |
instname_str |
Universidade Estadual Paulista (UNESP) |
instacron_str |
UNESP |
institution |
UNESP |
reponame_str |
Repositório Institucional da UNESP |
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Repositório Institucional da UNESP |
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Repositório Institucional da UNESP - Universidade Estadual Paulista (UNESP) |
repository.mail.fl_str_mv |
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1808128411677229056 |