Acetylcholinesterases from leaf-cutting ant atta sexdens: Purification, characterization, and capillary reactors for on-flow assays
Autor(a) principal: | |
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Data de Publicação: | 2019 |
Outros Autores: | , , , , , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Repositório Institucional da UNESP |
Texto Completo: | http://dx.doi.org/10.1155/2019/6139863 http://hdl.handle.net/11449/228701 |
Resumo: | Acetylcholinesterase (AChE) is responsible for catalyzing the hydrolysis of the neurotransmitter acetylcholine (ACh) leading to acetate and choline (Ch) release. The inhibition of AChE produces a generalized synaptic collapse that can lead to insect death. Herein we report for the first time the isolation of two AChEs from Atta sexdens which were purified by sulphate ammonium precipitation followed by ion exchange chromatography. AsAChE-A and AsAChE-B enzymes have optimum pH of 9.5 and 9.0 and higher activities in 30/50°C and 20°C, respectively, using acetylthiocholine (ATCh) as substrate. Immobilized capillary enzyme reactors (ICERs) were obtained for both enzymes (AsAChE-A-ICER and AsAChE-B-ICER) and their activities were measured by LC-MS/MS through hydrolysis product quantification of the natural substrate ACh. The comparison of activities by LC-MS/MS of both AChEs using ACh as substrate showed that AsAChE-B (free or immobilized) had the highest affinity. The inverse result was observed when the colorimetric assay (Elman method) was used for ATCh as substrate. Moreover, by mass spectrometry and phylogenetic studies, AsAChE-A and AsAChE-B were classified as belonging to AChE-2 and AChE-1 classes, respectively. |
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spelling |
Acetylcholinesterases from leaf-cutting ant atta sexdens: Purification, characterization, and capillary reactors for on-flow assaysAcetylcholinesterase (AChE) is responsible for catalyzing the hydrolysis of the neurotransmitter acetylcholine (ACh) leading to acetate and choline (Ch) release. The inhibition of AChE produces a generalized synaptic collapse that can lead to insect death. Herein we report for the first time the isolation of two AChEs from Atta sexdens which were purified by sulphate ammonium precipitation followed by ion exchange chromatography. AsAChE-A and AsAChE-B enzymes have optimum pH of 9.5 and 9.0 and higher activities in 30/50°C and 20°C, respectively, using acetylthiocholine (ATCh) as substrate. Immobilized capillary enzyme reactors (ICERs) were obtained for both enzymes (AsAChE-A-ICER and AsAChE-B-ICER) and their activities were measured by LC-MS/MS through hydrolysis product quantification of the natural substrate ACh. The comparison of activities by LC-MS/MS of both AChEs using ACh as substrate showed that AsAChE-B (free or immobilized) had the highest affinity. The inverse result was observed when the colorimetric assay (Elman method) was used for ATCh as substrate. Moreover, by mass spectrometry and phylogenetic studies, AsAChE-A and AsAChE-B were classified as belonging to AChE-2 and AChE-1 classes, respectively.Federal University of São Carlos Department of ChemistrySão Paulo University Instituto de Ciências Biomédicas (ICB)São Paulo State University Center for the Study of Social InsectsSão Paulo State University Center for the Study of Social InsectsUniversidade Federal de São Carlos (UFSCar)Universidade de São Paulo (USP)Universidade Estadual Paulista (UNESP)Dos Santos, Adriana M.Moreira, Ariele C.Lopes, Bianca RebeloFracola, Mariana F.De Almeida, Fernando G.Bueno, Odair C. [UNESP]Cass, Quezia B.Souza, Dulce Helena F.2022-04-29T08:28:18Z2022-04-29T08:28:18Z2019-01-01info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articlehttp://dx.doi.org/10.1155/2019/6139863Enzyme Research, v. 2019.2090-04142090-0406http://hdl.handle.net/11449/22870110.1155/2019/61398632-s2.0-85069050195Scopusreponame:Repositório Institucional da UNESPinstname:Universidade Estadual Paulista (UNESP)instacron:UNESPengEnzyme Researchinfo:eu-repo/semantics/openAccess2024-04-11T14:57:21Zoai:repositorio.unesp.br:11449/228701Repositório InstitucionalPUBhttp://repositorio.unesp.br/oai/requestopendoar:29462024-04-11T14:57:21Repositório Institucional da UNESP - Universidade Estadual Paulista (UNESP)false |
dc.title.none.fl_str_mv |
Acetylcholinesterases from leaf-cutting ant atta sexdens: Purification, characterization, and capillary reactors for on-flow assays |
title |
Acetylcholinesterases from leaf-cutting ant atta sexdens: Purification, characterization, and capillary reactors for on-flow assays |
spellingShingle |
Acetylcholinesterases from leaf-cutting ant atta sexdens: Purification, characterization, and capillary reactors for on-flow assays Dos Santos, Adriana M. |
title_short |
Acetylcholinesterases from leaf-cutting ant atta sexdens: Purification, characterization, and capillary reactors for on-flow assays |
title_full |
Acetylcholinesterases from leaf-cutting ant atta sexdens: Purification, characterization, and capillary reactors for on-flow assays |
title_fullStr |
Acetylcholinesterases from leaf-cutting ant atta sexdens: Purification, characterization, and capillary reactors for on-flow assays |
title_full_unstemmed |
Acetylcholinesterases from leaf-cutting ant atta sexdens: Purification, characterization, and capillary reactors for on-flow assays |
title_sort |
Acetylcholinesterases from leaf-cutting ant atta sexdens: Purification, characterization, and capillary reactors for on-flow assays |
author |
Dos Santos, Adriana M. |
author_facet |
Dos Santos, Adriana M. Moreira, Ariele C. Lopes, Bianca Rebelo Fracola, Mariana F. De Almeida, Fernando G. Bueno, Odair C. [UNESP] Cass, Quezia B. Souza, Dulce Helena F. |
author_role |
author |
author2 |
Moreira, Ariele C. Lopes, Bianca Rebelo Fracola, Mariana F. De Almeida, Fernando G. Bueno, Odair C. [UNESP] Cass, Quezia B. Souza, Dulce Helena F. |
author2_role |
author author author author author author author |
dc.contributor.none.fl_str_mv |
Universidade Federal de São Carlos (UFSCar) Universidade de São Paulo (USP) Universidade Estadual Paulista (UNESP) |
dc.contributor.author.fl_str_mv |
Dos Santos, Adriana M. Moreira, Ariele C. Lopes, Bianca Rebelo Fracola, Mariana F. De Almeida, Fernando G. Bueno, Odair C. [UNESP] Cass, Quezia B. Souza, Dulce Helena F. |
description |
Acetylcholinesterase (AChE) is responsible for catalyzing the hydrolysis of the neurotransmitter acetylcholine (ACh) leading to acetate and choline (Ch) release. The inhibition of AChE produces a generalized synaptic collapse that can lead to insect death. Herein we report for the first time the isolation of two AChEs from Atta sexdens which were purified by sulphate ammonium precipitation followed by ion exchange chromatography. AsAChE-A and AsAChE-B enzymes have optimum pH of 9.5 and 9.0 and higher activities in 30/50°C and 20°C, respectively, using acetylthiocholine (ATCh) as substrate. Immobilized capillary enzyme reactors (ICERs) were obtained for both enzymes (AsAChE-A-ICER and AsAChE-B-ICER) and their activities were measured by LC-MS/MS through hydrolysis product quantification of the natural substrate ACh. The comparison of activities by LC-MS/MS of both AChEs using ACh as substrate showed that AsAChE-B (free or immobilized) had the highest affinity. The inverse result was observed when the colorimetric assay (Elman method) was used for ATCh as substrate. Moreover, by mass spectrometry and phylogenetic studies, AsAChE-A and AsAChE-B were classified as belonging to AChE-2 and AChE-1 classes, respectively. |
publishDate |
2019 |
dc.date.none.fl_str_mv |
2019-01-01 2022-04-29T08:28:18Z 2022-04-29T08:28:18Z |
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/10.1155/2019/6139863 Enzyme Research, v. 2019. 2090-0414 2090-0406 http://hdl.handle.net/11449/228701 10.1155/2019/6139863 2-s2.0-85069050195 |
url |
http://dx.doi.org/10.1155/2019/6139863 http://hdl.handle.net/11449/228701 |
identifier_str_mv |
Enzyme Research, v. 2019. 2090-0414 2090-0406 10.1155/2019/6139863 2-s2.0-85069050195 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
Enzyme Research |
dc.rights.driver.fl_str_mv |
info:eu-repo/semantics/openAccess |
eu_rights_str_mv |
openAccess |
dc.source.none.fl_str_mv |
Scopus 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 |
collection |
Repositório Institucional da UNESP |
repository.name.fl_str_mv |
Repositório Institucional da UNESP - Universidade Estadual Paulista (UNESP) |
repository.mail.fl_str_mv |
|
_version_ |
1799965334514434048 |