High-level fed-batch fermentative expression of an engineered Staphylococcal protein A based ligand in E. coli: purification and characterization

Autores
Kangwa, Martin; Yelemane, Vikas; Polat, Ayse Nur; Gorrepati, Kanaka Durga Devi; Grasselli, Mariano; Fernández Lahore, Marcelo
Año de publicación
2015
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
The major platform for high level recombinant protein production is based on genetically modified microorganisms like Escherichia coli (E. coli) due to its short dividing time, ability to use inexpensive substrates and additionally, its genetics is comparatively simple, well characterized and can be manipulated easily. Here, we investigated the possibilities of finding the best media for high cell density fermentation, by analyzing different media samples, focusing on improving fermentation techniques and recombinant protein production. Initial fermentation of E. coli BL21 DE3:pAV01 in baffled flasks showed that high cell density was achieved when using complex media, Luria–Bertani (LB) and Terrific medium broth (TB) (10 and 14 g/L wet weight, respectively), as compared to mineral media M9, modified minimal medium (MMM) and Riesenberg mineral medium (RM) (7, 8 and 7 g/L, respectively). However, in fed-batch fermentation processes when using MMM after 25 h cultivation, it was possible to yield an optical density (OD600) of 139 corresponding to 172 g/L of wet biomass was produced in a 30 L TV Techfors-S Infors HT fermenter, with a computer controlled nutrient supply (glucose as a carbon source) delivery system, indicating nearly 1.5 times that obtained from TB. Upon purification, a total of 1.65 mg/g of protein per gram cell biomass was obtained and the purified AviPure showed affinity for immunoglobulin. High cell density fed batch fermentation was achieved by selecting the best media and growth conditions, by utilizing a number of fermentation parameters like media, fermentation conditions, chemical concentrations, pO2 level, stirrer speed, pH level and feed media addition. It is possible to reach cell densities higher than shake flasks and stirred tank reactors with the improved oxygen transfer rate and feed.
Fil: Kangwa, Martin. Jacobs University; Alemania
Fil: Yelemane, Vikas. Jacobs University; Alemania
Fil: Polat, Ayse Nur. Jacobs University; Alemania
Fil: Gorrepati, Kanaka Durga Devi. Jacobs University; Alemania
Fil: Grasselli, Mariano. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes; Argentina
Fil: Fernández Lahore, Marcelo. Jacobs University; Alemania
Materia
AVIPURE
E. COLI
FED-BATCH FERMENTATION
HIGH CELL DENSITY
RECOMBINANT PROTEIN A
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/53975
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/53975
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling High-level fed-batch fermentative expression of an engineered Staphylococcal protein A based ligand in E. coli: purification and characterizationKangwa, MartinYelemane, VikasPolat, Ayse NurGorrepati, Kanaka Durga DeviGrasselli, MarianoFernández Lahore, MarceloAVIPUREE. COLIFED-BATCH FERMENTATIONHIGH CELL DENSITYRECOMBINANT PROTEIN Ahttps://purl.org/becyt/ford/2.9https://purl.org/becyt/ford/2The major platform for high level recombinant protein production is based on genetically modified microorganisms like Escherichia coli (E. coli) due to its short dividing time, ability to use inexpensive substrates and additionally, its genetics is comparatively simple, well characterized and can be manipulated easily. Here, we investigated the possibilities of finding the best media for high cell density fermentation, by analyzing different media samples, focusing on improving fermentation techniques and recombinant protein production. Initial fermentation of E. coli BL21 DE3:pAV01 in baffled flasks showed that high cell density was achieved when using complex media, Luria–Bertani (LB) and Terrific medium broth (TB) (10 and 14 g/L wet weight, respectively), as compared to mineral media M9, modified minimal medium (MMM) and Riesenberg mineral medium (RM) (7, 8 and 7 g/L, respectively). However, in fed-batch fermentation processes when using MMM after 25 h cultivation, it was possible to yield an optical density (OD600) of 139 corresponding to 172 g/L of wet biomass was produced in a 30 L TV Techfors-S Infors HT fermenter, with a computer controlled nutrient supply (glucose as a carbon source) delivery system, indicating nearly 1.5 times that obtained from TB. Upon purification, a total of 1.65 mg/g of protein per gram cell biomass was obtained and the purified AviPure showed affinity for immunoglobulin. High cell density fed batch fermentation was achieved by selecting the best media and growth conditions, by utilizing a number of fermentation parameters like media, fermentation conditions, chemical concentrations, pO2 level, stirrer speed, pH level and feed media addition. It is possible to reach cell densities higher than shake flasks and stirred tank reactors with the improved oxygen transfer rate and feed.Fil: Kangwa, Martin. Jacobs University; AlemaniaFil: Yelemane, Vikas. Jacobs University; AlemaniaFil: Polat, Ayse Nur. Jacobs University; AlemaniaFil: Gorrepati, Kanaka Durga Devi. Jacobs University; AlemaniaFil: Grasselli, Mariano. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes; ArgentinaFil: Fernández Lahore, Marcelo. Jacobs University; AlemaniaSpringer Verlag2015-12info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/53975Kangwa, Martin; Yelemane, Vikas; Polat, Ayse Nur; Gorrepati, Kanaka Durga Devi; Grasselli, Mariano; et al.; High-level fed-batch fermentative expression of an engineered Staphylococcal protein A based ligand in E. coli: purification and characterization; Springer Verlag; AMB Express; 5; 1; 12-2015; 1-102191-0855CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/doi/10.1186/s13568-015-0155-yinfo:eu-repo/semantics/altIdentifier/url/https://amb-express.springeropen.com/articles/10.1186/s13568-015-0155-yinfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T09:33:18Zoai:ri.conicet.gov.ar:11336/53975instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-09-29 09:33:18.902CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv High-level fed-batch fermentative expression of an engineered Staphylococcal protein A based ligand in E. coli: purification and characterization
title High-level fed-batch fermentative expression of an engineered Staphylococcal protein A based ligand in E. coli: purification and characterization
spellingShingle High-level fed-batch fermentative expression of an engineered Staphylococcal protein A based ligand in E. coli: purification and characterization
Kangwa, Martin
AVIPURE
E. COLI
FED-BATCH FERMENTATION
HIGH CELL DENSITY
RECOMBINANT PROTEIN A
title_short High-level fed-batch fermentative expression of an engineered Staphylococcal protein A based ligand in E. coli: purification and characterization
title_full High-level fed-batch fermentative expression of an engineered Staphylococcal protein A based ligand in E. coli: purification and characterization
title_fullStr High-level fed-batch fermentative expression of an engineered Staphylococcal protein A based ligand in E. coli: purification and characterization
title_full_unstemmed High-level fed-batch fermentative expression of an engineered Staphylococcal protein A based ligand in E. coli: purification and characterization
title_sort High-level fed-batch fermentative expression of an engineered Staphylococcal protein A based ligand in E. coli: purification and characterization
dc.creator.none.fl_str_mv Kangwa, Martin
Yelemane, Vikas
Polat, Ayse Nur
Gorrepati, Kanaka Durga Devi
Grasselli, Mariano
Fernández Lahore, Marcelo
author Kangwa, Martin
author_facet Kangwa, Martin
Yelemane, Vikas
Polat, Ayse Nur
Gorrepati, Kanaka Durga Devi
Grasselli, Mariano
Fernández Lahore, Marcelo
author_role author
author2 Yelemane, Vikas
Polat, Ayse Nur
Gorrepati, Kanaka Durga Devi
Grasselli, Mariano
Fernández Lahore, Marcelo
author2_role author
author
author
author
author
dc.subject.none.fl_str_mv AVIPURE
E. COLI
FED-BATCH FERMENTATION
HIGH CELL DENSITY
RECOMBINANT PROTEIN A
topic AVIPURE
E. COLI
FED-BATCH FERMENTATION
HIGH CELL DENSITY
RECOMBINANT PROTEIN A
purl_subject.fl_str_mv https://purl.org/becyt/ford/2.9
https://purl.org/becyt/ford/2
dc.description.none.fl_txt_mv The major platform for high level recombinant protein production is based on genetically modified microorganisms like Escherichia coli (E. coli) due to its short dividing time, ability to use inexpensive substrates and additionally, its genetics is comparatively simple, well characterized and can be manipulated easily. Here, we investigated the possibilities of finding the best media for high cell density fermentation, by analyzing different media samples, focusing on improving fermentation techniques and recombinant protein production. Initial fermentation of E. coli BL21 DE3:pAV01 in baffled flasks showed that high cell density was achieved when using complex media, Luria–Bertani (LB) and Terrific medium broth (TB) (10 and 14 g/L wet weight, respectively), as compared to mineral media M9, modified minimal medium (MMM) and Riesenberg mineral medium (RM) (7, 8 and 7 g/L, respectively). However, in fed-batch fermentation processes when using MMM after 25 h cultivation, it was possible to yield an optical density (OD600) of 139 corresponding to 172 g/L of wet biomass was produced in a 30 L TV Techfors-S Infors HT fermenter, with a computer controlled nutrient supply (glucose as a carbon source) delivery system, indicating nearly 1.5 times that obtained from TB. Upon purification, a total of 1.65 mg/g of protein per gram cell biomass was obtained and the purified AviPure showed affinity for immunoglobulin. High cell density fed batch fermentation was achieved by selecting the best media and growth conditions, by utilizing a number of fermentation parameters like media, fermentation conditions, chemical concentrations, pO2 level, stirrer speed, pH level and feed media addition. It is possible to reach cell densities higher than shake flasks and stirred tank reactors with the improved oxygen transfer rate and feed.
Fil: Kangwa, Martin. Jacobs University; Alemania
Fil: Yelemane, Vikas. Jacobs University; Alemania
Fil: Polat, Ayse Nur. Jacobs University; Alemania
Fil: Gorrepati, Kanaka Durga Devi. Jacobs University; Alemania
Fil: Grasselli, Mariano. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes; Argentina
Fil: Fernández Lahore, Marcelo. Jacobs University; Alemania
description The major platform for high level recombinant protein production is based on genetically modified microorganisms like Escherichia coli (E. coli) due to its short dividing time, ability to use inexpensive substrates and additionally, its genetics is comparatively simple, well characterized and can be manipulated easily. Here, we investigated the possibilities of finding the best media for high cell density fermentation, by analyzing different media samples, focusing on improving fermentation techniques and recombinant protein production. Initial fermentation of E. coli BL21 DE3:pAV01 in baffled flasks showed that high cell density was achieved when using complex media, Luria–Bertani (LB) and Terrific medium broth (TB) (10 and 14 g/L wet weight, respectively), as compared to mineral media M9, modified minimal medium (MMM) and Riesenberg mineral medium (RM) (7, 8 and 7 g/L, respectively). However, in fed-batch fermentation processes when using MMM after 25 h cultivation, it was possible to yield an optical density (OD600) of 139 corresponding to 172 g/L of wet biomass was produced in a 30 L TV Techfors-S Infors HT fermenter, with a computer controlled nutrient supply (glucose as a carbon source) delivery system, indicating nearly 1.5 times that obtained from TB. Upon purification, a total of 1.65 mg/g of protein per gram cell biomass was obtained and the purified AviPure showed affinity for immunoglobulin. High cell density fed batch fermentation was achieved by selecting the best media and growth conditions, by utilizing a number of fermentation parameters like media, fermentation conditions, chemical concentrations, pO2 level, stirrer speed, pH level and feed media addition. It is possible to reach cell densities higher than shake flasks and stirred tank reactors with the improved oxygen transfer rate and feed.
publishDate 2015
dc.date.none.fl_str_mv 2015-12
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/53975
Kangwa, Martin; Yelemane, Vikas; Polat, Ayse Nur; Gorrepati, Kanaka Durga Devi; Grasselli, Mariano; et al.; High-level fed-batch fermentative expression of an engineered Staphylococcal protein A based ligand in E. coli: purification and characterization; Springer Verlag; AMB Express; 5; 1; 12-2015; 1-10
2191-0855
CONICET Digital
CONICET
url http://hdl.handle.net/11336/53975
identifier_str_mv Kangwa, Martin; Yelemane, Vikas; Polat, Ayse Nur; Gorrepati, Kanaka Durga Devi; Grasselli, Mariano; et al.; High-level fed-batch fermentative expression of an engineered Staphylococcal protein A based ligand in E. coli: purification and characterization; Springer Verlag; AMB Express; 5; 1; 12-2015; 1-10
2191-0855
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.1186/s13568-015-0155-y
info:eu-repo/semantics/altIdentifier/url/https://amb-express.springeropen.com/articles/10.1186/s13568-015-0155-y
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
dc.publisher.none.fl_str_mv Springer Verlag
publisher.none.fl_str_mv Springer Verlag
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
collection CONICET Digital (CONICET)
instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
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score 13.070432

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