Adaptation of the binding domain of Lactobacillus acidophilus S-layer protein as a molecular tag for affinity chromatography development

Autores
Muruaga, Emanuel Javier; Uriza, Paula Jimena; Eckert, Gonzalo Axel Klaus; Pepe, María Victoria; Duarte, Cecilia Magalí; Roset, Mara Sabrina; Briones, Carlos Gabriel
Año de publicación
2023
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Introduction: The S-layer proteins are a class of self-assembling proteins that form bi-dimensional lattices named S-Layer on the cell surface of bacteria and archaea. The protein SlpA, which is the major constituent of the Lactobacillus acidophilus S-layer, contains in its C-terminus region (SlpA284 − 444), a protein domain (named here as SLAPTAG) responsible for the association of SlpA to the bacterial surface. SLAPTAG was adapted for the development of a novel affinity chromatography method: the SLAPTAG-based affinity chromatography (SAC). Methods: Proteins with different molecular weights or biochemical functions were fused in-frame to the SLAPTAG and efficiently purified by a Bacillus subtilis-derived affinity matrix (named Bio-Matrix or BM). Different binding and elution conditions were evaluated to establish an optimized protocol. Results: The binding equilibrium between SLAPTAG and BM was reached after a few minutes of incubation at 4°C, with an apparent dissociation constant (KD) of 4.3μM. A reporter protein (H6-GFP-SLAPTAG) was used to compare SAC protein purification efficiency against commercial immobilized metal affinity chromatography. No differences in protein purification performance were observed between the two methods. The stability and reusability of the BM were evaluated, and it was found that the matrix remained stable for more than a year. BM could be reused up to five times without a significant loss in performance. Additionally, the recovery of bound SLAP-tagged proteins was explored using proteolysis with a SLAP-tagged version of the HRV-3c protease (SLAPASE). This released the untagged GFP while the cut SLAPTAG and the SLAPASE were retained in the BM. As an alternative, iron nanoparticles were linked to the BM, resulting in BMmag. The BMmag was successfully adapted for a magnetic SAC, a technique with potential applications in high-throughput protein production and purification. Discussion: The SAC protocol can be adapted as a universal tool for the purification of recombinant proteins. Furthermore, the SAC protocol utilizes simple and low-cost reagents, making it suitable for in-house protein purification systems in laboratories worldwide. This enables the production of pure recombinant proteins for research, diagnosis, and the food industry.
Fil: Muruaga, Emanuel Javier. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Uriza, Paula Jimena. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Eckert, Gonzalo Axel Klaus. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Pepe, María Victoria. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Duarte, Cecilia Magalí. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Roset, Mara Sabrina. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Briones, Carlos Gabriel. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Materia
LACTOBACILLUS ACIDOPHILUS
SLAP TAG
AFFINITY CHROMATOGRAPHY
BIO-MATRIX
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/228918
Acceder
id CONICETDig_637e55a298860509feef36c5ff94a681
oai_identifier_str oai:ri.conicet.gov.ar:11336/228918
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Adaptation of the binding domain of Lactobacillus acidophilus S-layer protein as a molecular tag for affinity chromatography developmentMuruaga, Emanuel JavierUriza, Paula JimenaEckert, Gonzalo Axel KlausPepe, María VictoriaDuarte, Cecilia MagalíRoset, Mara SabrinaBriones, Carlos GabrielLACTOBACILLUS ACIDOPHILUSSLAP TAGAFFINITY CHROMATOGRAPHYBIO-MATRIXhttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Introduction: The S-layer proteins are a class of self-assembling proteins that form bi-dimensional lattices named S-Layer on the cell surface of bacteria and archaea. The protein SlpA, which is the major constituent of the Lactobacillus acidophilus S-layer, contains in its C-terminus region (SlpA284 − 444), a protein domain (named here as SLAPTAG) responsible for the association of SlpA to the bacterial surface. SLAPTAG was adapted for the development of a novel affinity chromatography method: the SLAPTAG-based affinity chromatography (SAC). Methods: Proteins with different molecular weights or biochemical functions were fused in-frame to the SLAPTAG and efficiently purified by a Bacillus subtilis-derived affinity matrix (named Bio-Matrix or BM). Different binding and elution conditions were evaluated to establish an optimized protocol. Results: The binding equilibrium between SLAPTAG and BM was reached after a few minutes of incubation at 4°C, with an apparent dissociation constant (KD) of 4.3μM. A reporter protein (H6-GFP-SLAPTAG) was used to compare SAC protein purification efficiency against commercial immobilized metal affinity chromatography. No differences in protein purification performance were observed between the two methods. The stability and reusability of the BM were evaluated, and it was found that the matrix remained stable for more than a year. BM could be reused up to five times without a significant loss in performance. Additionally, the recovery of bound SLAP-tagged proteins was explored using proteolysis with a SLAP-tagged version of the HRV-3c protease (SLAPASE). This released the untagged GFP while the cut SLAPTAG and the SLAPASE were retained in the BM. As an alternative, iron nanoparticles were linked to the BM, resulting in BMmag. The BMmag was successfully adapted for a magnetic SAC, a technique with potential applications in high-throughput protein production and purification. Discussion: The SAC protocol can be adapted as a universal tool for the purification of recombinant proteins. Furthermore, the SAC protocol utilizes simple and low-cost reagents, making it suitable for in-house protein purification systems in laboratories worldwide. This enables the production of pure recombinant proteins for research, diagnosis, and the food industry.Fil: Muruaga, Emanuel Javier. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Uriza, Paula Jimena. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Eckert, Gonzalo Axel Klaus. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Pepe, María Victoria. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Duarte, Cecilia Magalí. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Roset, Mara Sabrina. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Briones, Carlos Gabriel. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; ArgentinaFrontiers Media2023-06info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/228918Muruaga, Emanuel Javier; Uriza, Paula Jimena; Eckert, Gonzalo Axel Klaus; Pepe, María Victoria; Duarte, Cecilia Magalí; et al.; Adaptation of the binding domain of Lactobacillus acidophilus S-layer protein as a molecular tag for affinity chromatography development; Frontiers Media; Frontiers in Microbiology; 14; 1210898; 6-2023; 1-151664-302XCONICET DigitalCONICETenghttps://ri.conicet.gov.ar/handle/11336/215848info:eu-repo/semantics/altIdentifier/url/https://www.frontiersin.org/articles/10.3389/fmicb.2023.1210898/fullinfo:eu-repo/semantics/altIdentifier/doi/10.3389/fmicb.2023.1210898info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-10-22T12:14:13Zoai:ri.conicet.gov.ar:11336/228918instacron: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:34982025-10-22 12:14:13.546CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Adaptation of the binding domain of Lactobacillus acidophilus S-layer protein as a molecular tag for affinity chromatography development
title Adaptation of the binding domain of Lactobacillus acidophilus S-layer protein as a molecular tag for affinity chromatography development
spellingShingle Adaptation of the binding domain of Lactobacillus acidophilus S-layer protein as a molecular tag for affinity chromatography development
Muruaga, Emanuel Javier
LACTOBACILLUS ACIDOPHILUS
SLAP TAG
AFFINITY CHROMATOGRAPHY
BIO-MATRIX
title_short Adaptation of the binding domain of Lactobacillus acidophilus S-layer protein as a molecular tag for affinity chromatography development
title_full Adaptation of the binding domain of Lactobacillus acidophilus S-layer protein as a molecular tag for affinity chromatography development
title_fullStr Adaptation of the binding domain of Lactobacillus acidophilus S-layer protein as a molecular tag for affinity chromatography development
title_full_unstemmed Adaptation of the binding domain of Lactobacillus acidophilus S-layer protein as a molecular tag for affinity chromatography development
title_sort Adaptation of the binding domain of Lactobacillus acidophilus S-layer protein as a molecular tag for affinity chromatography development
dc.creator.none.fl_str_mv Muruaga, Emanuel Javier
Uriza, Paula Jimena
Eckert, Gonzalo Axel Klaus
Pepe, María Victoria
Duarte, Cecilia Magalí
Roset, Mara Sabrina
Briones, Carlos Gabriel
author Muruaga, Emanuel Javier
author_facet Muruaga, Emanuel Javier
Uriza, Paula Jimena
Eckert, Gonzalo Axel Klaus
Pepe, María Victoria
Duarte, Cecilia Magalí
Roset, Mara Sabrina
Briones, Carlos Gabriel
author_role author
author2 Uriza, Paula Jimena
Eckert, Gonzalo Axel Klaus
Pepe, María Victoria
Duarte, Cecilia Magalí
Roset, Mara Sabrina
Briones, Carlos Gabriel
author2_role author
author
author
author
author
author
dc.subject.none.fl_str_mv LACTOBACILLUS ACIDOPHILUS
SLAP TAG
AFFINITY CHROMATOGRAPHY
BIO-MATRIX
topic LACTOBACILLUS ACIDOPHILUS
SLAP TAG
AFFINITY CHROMATOGRAPHY
BIO-MATRIX
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv Introduction: The S-layer proteins are a class of self-assembling proteins that form bi-dimensional lattices named S-Layer on the cell surface of bacteria and archaea. The protein SlpA, which is the major constituent of the Lactobacillus acidophilus S-layer, contains in its C-terminus region (SlpA284 − 444), a protein domain (named here as SLAPTAG) responsible for the association of SlpA to the bacterial surface. SLAPTAG was adapted for the development of a novel affinity chromatography method: the SLAPTAG-based affinity chromatography (SAC). Methods: Proteins with different molecular weights or biochemical functions were fused in-frame to the SLAPTAG and efficiently purified by a Bacillus subtilis-derived affinity matrix (named Bio-Matrix or BM). Different binding and elution conditions were evaluated to establish an optimized protocol. Results: The binding equilibrium between SLAPTAG and BM was reached after a few minutes of incubation at 4°C, with an apparent dissociation constant (KD) of 4.3μM. A reporter protein (H6-GFP-SLAPTAG) was used to compare SAC protein purification efficiency against commercial immobilized metal affinity chromatography. No differences in protein purification performance were observed between the two methods. The stability and reusability of the BM were evaluated, and it was found that the matrix remained stable for more than a year. BM could be reused up to five times without a significant loss in performance. Additionally, the recovery of bound SLAP-tagged proteins was explored using proteolysis with a SLAP-tagged version of the HRV-3c protease (SLAPASE). This released the untagged GFP while the cut SLAPTAG and the SLAPASE were retained in the BM. As an alternative, iron nanoparticles were linked to the BM, resulting in BMmag. The BMmag was successfully adapted for a magnetic SAC, a technique with potential applications in high-throughput protein production and purification. Discussion: The SAC protocol can be adapted as a universal tool for the purification of recombinant proteins. Furthermore, the SAC protocol utilizes simple and low-cost reagents, making it suitable for in-house protein purification systems in laboratories worldwide. This enables the production of pure recombinant proteins for research, diagnosis, and the food industry.
Fil: Muruaga, Emanuel Javier. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Uriza, Paula Jimena. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Eckert, Gonzalo Axel Klaus. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Pepe, María Victoria. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Duarte, Cecilia Magalí. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Roset, Mara Sabrina. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Briones, Carlos Gabriel. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
description Introduction: The S-layer proteins are a class of self-assembling proteins that form bi-dimensional lattices named S-Layer on the cell surface of bacteria and archaea. The protein SlpA, which is the major constituent of the Lactobacillus acidophilus S-layer, contains in its C-terminus region (SlpA284 − 444), a protein domain (named here as SLAPTAG) responsible for the association of SlpA to the bacterial surface. SLAPTAG was adapted for the development of a novel affinity chromatography method: the SLAPTAG-based affinity chromatography (SAC). Methods: Proteins with different molecular weights or biochemical functions were fused in-frame to the SLAPTAG and efficiently purified by a Bacillus subtilis-derived affinity matrix (named Bio-Matrix or BM). Different binding and elution conditions were evaluated to establish an optimized protocol. Results: The binding equilibrium between SLAPTAG and BM was reached after a few minutes of incubation at 4°C, with an apparent dissociation constant (KD) of 4.3μM. A reporter protein (H6-GFP-SLAPTAG) was used to compare SAC protein purification efficiency against commercial immobilized metal affinity chromatography. No differences in protein purification performance were observed between the two methods. The stability and reusability of the BM were evaluated, and it was found that the matrix remained stable for more than a year. BM could be reused up to five times without a significant loss in performance. Additionally, the recovery of bound SLAP-tagged proteins was explored using proteolysis with a SLAP-tagged version of the HRV-3c protease (SLAPASE). This released the untagged GFP while the cut SLAPTAG and the SLAPASE were retained in the BM. As an alternative, iron nanoparticles were linked to the BM, resulting in BMmag. The BMmag was successfully adapted for a magnetic SAC, a technique with potential applications in high-throughput protein production and purification. Discussion: The SAC protocol can be adapted as a universal tool for the purification of recombinant proteins. Furthermore, the SAC protocol utilizes simple and low-cost reagents, making it suitable for in-house protein purification systems in laboratories worldwide. This enables the production of pure recombinant proteins for research, diagnosis, and the food industry.
publishDate 2023
dc.date.none.fl_str_mv 2023-06
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/11336/228918
Muruaga, Emanuel Javier; Uriza, Paula Jimena; Eckert, Gonzalo Axel Klaus; Pepe, María Victoria; Duarte, Cecilia Magalí; et al.; Adaptation of the binding domain of Lactobacillus acidophilus S-layer protein as a molecular tag for affinity chromatography development; Frontiers Media; Frontiers in Microbiology; 14; 1210898; 6-2023; 1-15
1664-302X
CONICET Digital
CONICET
url http://hdl.handle.net/11336/228918
identifier_str_mv Muruaga, Emanuel Javier; Uriza, Paula Jimena; Eckert, Gonzalo Axel Klaus; Pepe, María Victoria; Duarte, Cecilia Magalí; et al.; Adaptation of the binding domain of Lactobacillus acidophilus S-layer protein as a molecular tag for affinity chromatography development; Frontiers Media; Frontiers in Microbiology; 14; 1210898; 6-2023; 1-15
1664-302X
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv https://ri.conicet.gov.ar/handle/11336/215848
info:eu-repo/semantics/altIdentifier/url/https://www.frontiersin.org/articles/10.3389/fmicb.2023.1210898/full
info:eu-repo/semantics/altIdentifier/doi/10.3389/fmicb.2023.1210898
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
application/pdf
application/pdf
dc.publisher.none.fl_str_mv Frontiers Media
publisher.none.fl_str_mv Frontiers Media
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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