Cloning and characterization of the iron uptake gene iutA from avian Escherichia coli
Autor(a) principal: | |
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Data de Publicação: | 2008 |
Outros Autores: | , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Brazilian Archives of Biology and Technology |
Texto Completo: | http://old.scielo.br/scielo.php?script=sci_arttext&pid=S1516-89132008000300006 |
Resumo: | The aim of this work was to isolate, clone and characterize the iron uptake gene iutA from avian pathogenic E. coli (APEC). The iutA gene was isolated from the strain APEC 9, serotype O2:H9, which was cloned in the expression vector pET101/D-TOPO. The gene of 2.2 Kb was sequenced (AY602767, which showed high similarity to the iutA gene from three plasmids, two from APEC, pAPEC-02-ColV (AY545598.4) and pTJ100 (AY553855.1), and one from a human invasive E. coli strain, the pColV K30. The recombinant protein IutA was over expressed in E. coli BL21(DE-3) and was solubilized with urea and purified by Ni-NTA column. This method produced a relatively high yield of r-IutA of approximately 74kDa, which was used to produce the antibody anti-IutA. This anti-IutA reacted with the protein r-IutA and native IutA of APEC 9, as demonstrated by Western blot, showing that the r-IutA conserved epitopes and its antigenicity was preserved. The anti-IutA IgY was able to inhibit the IutA biological activity, inhibiting the sensitivity to cloacin DF13 of APEC9. However, it did not inhibit the growth of APEC9 in M9 and did not protect the chickens inoculated with the APEC, suggesting that the APEC possessed another iron acquisition mechanism distinct of aerobactin. |
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Cloning and characterization of the iron uptake gene iutA from avian Escherichia coliAvian Escherichia coliiron uptakeaerobactin receptorvirulence factoriutA geneThe aim of this work was to isolate, clone and characterize the iron uptake gene iutA from avian pathogenic E. coli (APEC). The iutA gene was isolated from the strain APEC 9, serotype O2:H9, which was cloned in the expression vector pET101/D-TOPO. The gene of 2.2 Kb was sequenced (AY602767, which showed high similarity to the iutA gene from three plasmids, two from APEC, pAPEC-02-ColV (AY545598.4) and pTJ100 (AY553855.1), and one from a human invasive E. coli strain, the pColV K30. The recombinant protein IutA was over expressed in E. coli BL21(DE-3) and was solubilized with urea and purified by Ni-NTA column. This method produced a relatively high yield of r-IutA of approximately 74kDa, which was used to produce the antibody anti-IutA. This anti-IutA reacted with the protein r-IutA and native IutA of APEC 9, as demonstrated by Western blot, showing that the r-IutA conserved epitopes and its antigenicity was preserved. The anti-IutA IgY was able to inhibit the IutA biological activity, inhibiting the sensitivity to cloacin DF13 of APEC9. However, it did not inhibit the growth of APEC9 in M9 and did not protect the chickens inoculated with the APEC, suggesting that the APEC possessed another iron acquisition mechanism distinct of aerobactin.Instituto de Tecnologia do Paraná - Tecpar2008-06-01info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersiontext/htmlhttp://old.scielo.br/scielo.php?script=sci_arttext&pid=S1516-89132008000300006Brazilian Archives of Biology and Technology v.51 n.3 2008reponame:Brazilian Archives of Biology and Technologyinstname:Instituto de Tecnologia do Paraná (Tecpar)instacron:TECPAR10.1590/S1516-89132008000300006info:eu-repo/semantics/openAccessTokano,Dorismey VieiraKawaichi,Marisa EmikoVenâncio,Emerson JoséVidotto,Marilda Carloseng2008-07-16T00:00:00Zoai:scielo:S1516-89132008000300006Revistahttps://www.scielo.br/j/babt/https://old.scielo.br/oai/scielo-oai.phpbabt@tecpar.br||babt@tecpar.br1678-43241516-8913opendoar:2008-07-16T00:00Brazilian Archives of Biology and Technology - Instituto de Tecnologia do Paraná (Tecpar)false |
dc.title.none.fl_str_mv |
Cloning and characterization of the iron uptake gene iutA from avian Escherichia coli |
title |
Cloning and characterization of the iron uptake gene iutA from avian Escherichia coli |
spellingShingle |
Cloning and characterization of the iron uptake gene iutA from avian Escherichia coli Tokano,Dorismey Vieira Avian Escherichia coli iron uptake aerobactin receptor virulence factor iutA gene |
title_short |
Cloning and characterization of the iron uptake gene iutA from avian Escherichia coli |
title_full |
Cloning and characterization of the iron uptake gene iutA from avian Escherichia coli |
title_fullStr |
Cloning and characterization of the iron uptake gene iutA from avian Escherichia coli |
title_full_unstemmed |
Cloning and characterization of the iron uptake gene iutA from avian Escherichia coli |
title_sort |
Cloning and characterization of the iron uptake gene iutA from avian Escherichia coli |
author |
Tokano,Dorismey Vieira |
author_facet |
Tokano,Dorismey Vieira Kawaichi,Marisa Emiko Venâncio,Emerson José Vidotto,Marilda Carlos |
author_role |
author |
author2 |
Kawaichi,Marisa Emiko Venâncio,Emerson José Vidotto,Marilda Carlos |
author2_role |
author author author |
dc.contributor.author.fl_str_mv |
Tokano,Dorismey Vieira Kawaichi,Marisa Emiko Venâncio,Emerson José Vidotto,Marilda Carlos |
dc.subject.por.fl_str_mv |
Avian Escherichia coli iron uptake aerobactin receptor virulence factor iutA gene |
topic |
Avian Escherichia coli iron uptake aerobactin receptor virulence factor iutA gene |
description |
The aim of this work was to isolate, clone and characterize the iron uptake gene iutA from avian pathogenic E. coli (APEC). The iutA gene was isolated from the strain APEC 9, serotype O2:H9, which was cloned in the expression vector pET101/D-TOPO. The gene of 2.2 Kb was sequenced (AY602767, which showed high similarity to the iutA gene from three plasmids, two from APEC, pAPEC-02-ColV (AY545598.4) and pTJ100 (AY553855.1), and one from a human invasive E. coli strain, the pColV K30. The recombinant protein IutA was over expressed in E. coli BL21(DE-3) and was solubilized with urea and purified by Ni-NTA column. This method produced a relatively high yield of r-IutA of approximately 74kDa, which was used to produce the antibody anti-IutA. This anti-IutA reacted with the protein r-IutA and native IutA of APEC 9, as demonstrated by Western blot, showing that the r-IutA conserved epitopes and its antigenicity was preserved. The anti-IutA IgY was able to inhibit the IutA biological activity, inhibiting the sensitivity to cloacin DF13 of APEC9. However, it did not inhibit the growth of APEC9 in M9 and did not protect the chickens inoculated with the APEC, suggesting that the APEC possessed another iron acquisition mechanism distinct of aerobactin. |
publishDate |
2008 |
dc.date.none.fl_str_mv |
2008-06-01 |
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=S1516-89132008000300006 |
url |
http://old.scielo.br/scielo.php?script=sci_arttext&pid=S1516-89132008000300006 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
10.1590/S1516-89132008000300006 |
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 |
Instituto de Tecnologia do Paraná - Tecpar |
publisher.none.fl_str_mv |
Instituto de Tecnologia do Paraná - Tecpar |
dc.source.none.fl_str_mv |
Brazilian Archives of Biology and Technology v.51 n.3 2008 reponame:Brazilian Archives of Biology and Technology instname:Instituto de Tecnologia do Paraná (Tecpar) instacron:TECPAR |
instname_str |
Instituto de Tecnologia do Paraná (Tecpar) |
instacron_str |
TECPAR |
institution |
TECPAR |
reponame_str |
Brazilian Archives of Biology and Technology |
collection |
Brazilian Archives of Biology and Technology |
repository.name.fl_str_mv |
Brazilian Archives of Biology and Technology - Instituto de Tecnologia do Paraná (Tecpar) |
repository.mail.fl_str_mv |
babt@tecpar.br||babt@tecpar.br |
_version_ |
1750318272244350976 |