Determination of disulfide bridges of two spider toxins: hainantoxin-III and hainantoxin-IV
Autor(a) principal: | |
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Data de Publicação: | 2009 |
Outros Autores: | , , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | The Journal of venomous animals and toxins including tropical diseases (Online) |
Texto Completo: | http://old.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992009000200009 |
Resumo: | Peptide toxins are usually highly bridged proteins with multipairs of intrachain disulfide bonds. Analysis of disulfide connectivity is an important facet of protein structure determination. In this paper, we successfully assigned the disulfide linkage of two novel peptide toxins, called HNTX-III and HNTX-IV, isolated from the venom of Ornithoctonus hainana spider. Both peptides are useful inhibitors of TTX-sensitive voltage-gated sodium channels and are composed of six cysteine residues that form three disulfide bonds, respectively. Firstly, the peptides were partially reduced by tris(2-carboxyethyl)-phosphine (TCEP) in 0.1 M citrate buffer containing 6 M guanidine-HCl at 40° C for ten minutes. Subsequently, the partially reduced intermediates containing free thiols were separated by reversed-phase high-performance liquid chromatography (RP-HPLC) and alkylated by rapid carboxamidomethylation. Then, the disulfide bonds of the intermediates were analyzed by Edman degradation. By using the strategy above, disulfide linkages of HNTX-III and HNTX-IV were determined as I-IV, II-V and III-VI pattern. In addition, this study also showed that this method may have a great potential for determining the disulfide bonds of spider peptide toxins. |
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The Journal of venomous animals and toxins including tropical diseases (Online) |
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Determination of disulfide bridges of two spider toxins: hainantoxin-III and hainantoxin-IVdisulfide bondsTCEPpartial reductionHNTX-IIIHNTX-IVPeptide toxins are usually highly bridged proteins with multipairs of intrachain disulfide bonds. Analysis of disulfide connectivity is an important facet of protein structure determination. In this paper, we successfully assigned the disulfide linkage of two novel peptide toxins, called HNTX-III and HNTX-IV, isolated from the venom of Ornithoctonus hainana spider. Both peptides are useful inhibitors of TTX-sensitive voltage-gated sodium channels and are composed of six cysteine residues that form three disulfide bonds, respectively. Firstly, the peptides were partially reduced by tris(2-carboxyethyl)-phosphine (TCEP) in 0.1 M citrate buffer containing 6 M guanidine-HCl at 40° C for ten minutes. Subsequently, the partially reduced intermediates containing free thiols were separated by reversed-phase high-performance liquid chromatography (RP-HPLC) and alkylated by rapid carboxamidomethylation. Then, the disulfide bonds of the intermediates were analyzed by Edman degradation. By using the strategy above, disulfide linkages of HNTX-III and HNTX-IV were determined as I-IV, II-V and III-VI pattern. In addition, this study also showed that this method may have a great potential for determining the disulfide bonds of spider peptide toxins.Centro de Estudos de Venenos e Animais Peçonhentos (CEVAP/UNESP)2009-01-01info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersiontext/htmlhttp://old.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992009000200009Journal of Venomous Animals and Toxins including Tropical Diseases v.15 n.2 2009reponame:The Journal of venomous animals and toxins including tropical diseases (Online)instname:Universidade Estadual Paulista (UNESP)instacron:UNESP10.1590/S1678-91992009000200009info:eu-repo/semantics/openAccessWang,WLiu,ZQian,WFang,YLiang,Seng2009-06-09T00:00:00Zoai:scielo:S1678-91992009000200009Revistahttp://www.scielo.br/jvatitdPUBhttps://old.scielo.br/oai/scielo-oai.php||editorial@jvat.org.br1678-91991678-9180opendoar:2009-06-09T00:00The Journal of venomous animals and toxins including tropical diseases (Online) - Universidade Estadual Paulista (UNESP)false |
dc.title.none.fl_str_mv |
Determination of disulfide bridges of two spider toxins: hainantoxin-III and hainantoxin-IV |
title |
Determination of disulfide bridges of two spider toxins: hainantoxin-III and hainantoxin-IV |
spellingShingle |
Determination of disulfide bridges of two spider toxins: hainantoxin-III and hainantoxin-IV Wang,W disulfide bonds TCEP partial reduction HNTX-III HNTX-IV |
title_short |
Determination of disulfide bridges of two spider toxins: hainantoxin-III and hainantoxin-IV |
title_full |
Determination of disulfide bridges of two spider toxins: hainantoxin-III and hainantoxin-IV |
title_fullStr |
Determination of disulfide bridges of two spider toxins: hainantoxin-III and hainantoxin-IV |
title_full_unstemmed |
Determination of disulfide bridges of two spider toxins: hainantoxin-III and hainantoxin-IV |
title_sort |
Determination of disulfide bridges of two spider toxins: hainantoxin-III and hainantoxin-IV |
author |
Wang,W |
author_facet |
Wang,W Liu,Z Qian,W Fang,Y Liang,S |
author_role |
author |
author2 |
Liu,Z Qian,W Fang,Y Liang,S |
author2_role |
author author author author |
dc.contributor.author.fl_str_mv |
Wang,W Liu,Z Qian,W Fang,Y Liang,S |
dc.subject.por.fl_str_mv |
disulfide bonds TCEP partial reduction HNTX-III HNTX-IV |
topic |
disulfide bonds TCEP partial reduction HNTX-III HNTX-IV |
description |
Peptide toxins are usually highly bridged proteins with multipairs of intrachain disulfide bonds. Analysis of disulfide connectivity is an important facet of protein structure determination. In this paper, we successfully assigned the disulfide linkage of two novel peptide toxins, called HNTX-III and HNTX-IV, isolated from the venom of Ornithoctonus hainana spider. Both peptides are useful inhibitors of TTX-sensitive voltage-gated sodium channels and are composed of six cysteine residues that form three disulfide bonds, respectively. Firstly, the peptides were partially reduced by tris(2-carboxyethyl)-phosphine (TCEP) in 0.1 M citrate buffer containing 6 M guanidine-HCl at 40° C for ten minutes. Subsequently, the partially reduced intermediates containing free thiols were separated by reversed-phase high-performance liquid chromatography (RP-HPLC) and alkylated by rapid carboxamidomethylation. Then, the disulfide bonds of the intermediates were analyzed by Edman degradation. By using the strategy above, disulfide linkages of HNTX-III and HNTX-IV were determined as I-IV, II-V and III-VI pattern. In addition, this study also showed that this method may have a great potential for determining the disulfide bonds of spider peptide toxins. |
publishDate |
2009 |
dc.date.none.fl_str_mv |
2009-01-01 |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/article |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
format |
article |
status_str |
publishedVersion |
dc.identifier.uri.fl_str_mv |
http://old.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992009000200009 |
url |
http://old.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992009000200009 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
10.1590/S1678-91992009000200009 |
dc.rights.driver.fl_str_mv |
info:eu-repo/semantics/openAccess |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
text/html |
dc.publisher.none.fl_str_mv |
Centro de Estudos de Venenos e Animais Peçonhentos (CEVAP/UNESP) |
publisher.none.fl_str_mv |
Centro de Estudos de Venenos e Animais Peçonhentos (CEVAP/UNESP) |
dc.source.none.fl_str_mv |
Journal of Venomous Animals and Toxins including Tropical Diseases v.15 n.2 2009 reponame:The Journal of venomous animals and toxins including tropical diseases (Online) instname:Universidade Estadual Paulista (UNESP) instacron:UNESP |
instname_str |
Universidade Estadual Paulista (UNESP) |
instacron_str |
UNESP |
institution |
UNESP |
reponame_str |
The Journal of venomous animals and toxins including tropical diseases (Online) |
collection |
The Journal of venomous animals and toxins including tropical diseases (Online) |
repository.name.fl_str_mv |
The Journal of venomous animals and toxins including tropical diseases (Online) - Universidade Estadual Paulista (UNESP) |
repository.mail.fl_str_mv |
||editorial@jvat.org.br |
_version_ |
1748958538397384704 |