Cloning, expression and characterization of alcohol dehydrogenases in the silkworm Bombyx mori
Autor(a) principal: | |
---|---|
Data de Publicação: | 2011 |
Outros Autores: | , , , , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Genetics and Molecular Biology |
Texto Completo: | http://old.scielo.br/scielo.php?script=sci_arttext&pid=S1415-47572011000200013 |
Resumo: | Alcohol dehydrogenases (ADH) are a class of enzymes that catalyze the reversible oxidation of alcohols to corresponding aldehydes or ketones, by using either nicotinamide adenine dinucleotide (NAD) or nicotinamide adenine dinucleotide phosphate (NADP), as coenzymes. In this study, a short-chain ADH gene was identified in Bombyx mori by 5'-RACE PCR. This is the first time the coding region of BmADH has been cloned, expressed, purified and then characterized. The cDNA fragment encoding the BmADH protein was amplified from a pool of silkworm cDNAs by PCR, and then cloned into E. coli expression vector pET-30a(+). The recombinant His-tagged BmADH protein was expressed in E. coli BL21 (DE3), and then purified by metal chelating affinity chromatography. The soluble recombinant BmADH, produced at low-growth temperature, was instrumental in catalyzing the ethanol-dependent reduction of NAD+, thereby indicating ethanol as one of the substrates of BmADH. |
id |
SBG-1_218dedcc216363acae7077ba2238406a |
---|---|
oai_identifier_str |
oai:scielo:S1415-47572011000200013 |
network_acronym_str |
SBG-1 |
network_name_str |
Genetics and Molecular Biology |
repository_id_str |
|
spelling |
Cloning, expression and characterization of alcohol dehydrogenases in the silkworm Bombyx mori5'-RACE PCRADHenzymatic activityrecombinant proteinAlcohol dehydrogenases (ADH) are a class of enzymes that catalyze the reversible oxidation of alcohols to corresponding aldehydes or ketones, by using either nicotinamide adenine dinucleotide (NAD) or nicotinamide adenine dinucleotide phosphate (NADP), as coenzymes. In this study, a short-chain ADH gene was identified in Bombyx mori by 5'-RACE PCR. This is the first time the coding region of BmADH has been cloned, expressed, purified and then characterized. The cDNA fragment encoding the BmADH protein was amplified from a pool of silkworm cDNAs by PCR, and then cloned into E. coli expression vector pET-30a(+). The recombinant His-tagged BmADH protein was expressed in E. coli BL21 (DE3), and then purified by metal chelating affinity chromatography. The soluble recombinant BmADH, produced at low-growth temperature, was instrumental in catalyzing the ethanol-dependent reduction of NAD+, thereby indicating ethanol as one of the substrates of BmADH.Sociedade Brasileira de Genética2011-01-01info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersiontext/htmlhttp://old.scielo.br/scielo.php?script=sci_arttext&pid=S1415-47572011000200013Genetics and Molecular Biology v.34 n.2 2011reponame:Genetics and Molecular Biologyinstname:Sociedade Brasileira de Genética (SBG)instacron:SBG10.1590/S1415-47572011000200013info:eu-repo/semantics/openAccessWang,NanShi,HaifengYao,QinZhou,YangKang,LequnChen,HuiqinChen,Kepingeng2011-06-02T00:00:00Zoai:scielo:S1415-47572011000200013Revistahttp://www.gmb.org.br/ONGhttps://old.scielo.br/oai/scielo-oai.php||editor@gmb.org.br1678-46851415-4757opendoar:2011-06-02T00:00Genetics and Molecular Biology - Sociedade Brasileira de Genética (SBG)false |
dc.title.none.fl_str_mv |
Cloning, expression and characterization of alcohol dehydrogenases in the silkworm Bombyx mori |
title |
Cloning, expression and characterization of alcohol dehydrogenases in the silkworm Bombyx mori |
spellingShingle |
Cloning, expression and characterization of alcohol dehydrogenases in the silkworm Bombyx mori Wang,Nan 5'-RACE PCRADH enzymatic activity recombinant protein |
title_short |
Cloning, expression and characterization of alcohol dehydrogenases in the silkworm Bombyx mori |
title_full |
Cloning, expression and characterization of alcohol dehydrogenases in the silkworm Bombyx mori |
title_fullStr |
Cloning, expression and characterization of alcohol dehydrogenases in the silkworm Bombyx mori |
title_full_unstemmed |
Cloning, expression and characterization of alcohol dehydrogenases in the silkworm Bombyx mori |
title_sort |
Cloning, expression and characterization of alcohol dehydrogenases in the silkworm Bombyx mori |
author |
Wang,Nan |
author_facet |
Wang,Nan Shi,Haifeng Yao,Qin Zhou,Yang Kang,Lequn Chen,Huiqin Chen,Keping |
author_role |
author |
author2 |
Shi,Haifeng Yao,Qin Zhou,Yang Kang,Lequn Chen,Huiqin Chen,Keping |
author2_role |
author author author author author author |
dc.contributor.author.fl_str_mv |
Wang,Nan Shi,Haifeng Yao,Qin Zhou,Yang Kang,Lequn Chen,Huiqin Chen,Keping |
dc.subject.por.fl_str_mv |
5'-RACE PCRADH enzymatic activity recombinant protein |
topic |
5'-RACE PCRADH enzymatic activity recombinant protein |
description |
Alcohol dehydrogenases (ADH) are a class of enzymes that catalyze the reversible oxidation of alcohols to corresponding aldehydes or ketones, by using either nicotinamide adenine dinucleotide (NAD) or nicotinamide adenine dinucleotide phosphate (NADP), as coenzymes. In this study, a short-chain ADH gene was identified in Bombyx mori by 5'-RACE PCR. This is the first time the coding region of BmADH has been cloned, expressed, purified and then characterized. The cDNA fragment encoding the BmADH protein was amplified from a pool of silkworm cDNAs by PCR, and then cloned into E. coli expression vector pET-30a(+). The recombinant His-tagged BmADH protein was expressed in E. coli BL21 (DE3), and then purified by metal chelating affinity chromatography. The soluble recombinant BmADH, produced at low-growth temperature, was instrumental in catalyzing the ethanol-dependent reduction of NAD+, thereby indicating ethanol as one of the substrates of BmADH. |
publishDate |
2011 |
dc.date.none.fl_str_mv |
2011-01-01 |
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=S1415-47572011000200013 |
url |
http://old.scielo.br/scielo.php?script=sci_arttext&pid=S1415-47572011000200013 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
10.1590/S1415-47572011000200013 |
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 |
Sociedade Brasileira de Genética |
publisher.none.fl_str_mv |
Sociedade Brasileira de Genética |
dc.source.none.fl_str_mv |
Genetics and Molecular Biology v.34 n.2 2011 reponame:Genetics and Molecular Biology instname:Sociedade Brasileira de Genética (SBG) instacron:SBG |
instname_str |
Sociedade Brasileira de Genética (SBG) |
instacron_str |
SBG |
institution |
SBG |
reponame_str |
Genetics and Molecular Biology |
collection |
Genetics and Molecular Biology |
repository.name.fl_str_mv |
Genetics and Molecular Biology - Sociedade Brasileira de Genética (SBG) |
repository.mail.fl_str_mv |
||editor@gmb.org.br |
_version_ |
1752122383717629952 |