Apolipoprotein E4 heterologous expression, purification under non-denaturing conditions, and effects on neuronal clonal cell lines

Abstract: The ε4 allele of the apolipoprotein E gene (APOE4) constitutes the main genetic risk factor for late-onset Alzheimer disease (AD). High amounts of pure apolipoprotein E4 (ApoE4), in a rapid and reproducible fashion, could be of value for studying its pathophysiological roles in AD. The aim...

Descripción completa

Guardado en:
Detalles Bibliográficos
Autores principales: Serrano, Ezequiel, Barrantes, Francisco José, Valdivieso, Ángel Gabriel
Formato: Artículo
Lenguaje:Español
Publicado: Elsevier 2023
Materias:
ENFERMEDAD DE ALZHEIMER
PROTEINAS
APOLIPOPROTEINA
Acceso en línea:https://repositorio.uca.edu.ar/handle/123456789/16628
Aporte de:
Repositorio Institucional de la Universidad Católica Argentina (UCA) de Universidad Católica Argentina
id I33-R139-123456789-16628
record_format dspace
spelling I33-R139-123456789-166282023-06-29T05:02:03Z Apolipoprotein E4 heterologous expression, purification under non-denaturing conditions, and effects on neuronal clonal cell lines Serrano, Ezequiel Barrantes, Francisco José Valdivieso, Ángel Gabriel ENFERMEDAD DE ALZHEIMER PROTEINAS APOLIPOPROTEINA Abstract: The ε4 allele of the apolipoprotein E gene (APOE4) constitutes the main genetic risk factor for late-onset Alzheimer disease (AD). High amounts of pure apolipoprotein E4 (ApoE4), in a rapid and reproducible fashion, could be of value for studying its pathophysiological roles in AD. The aim of the present work was to optimize a preparative method to obtain highly purified recombinant ApoE4 (rApoE4) with full biological activity. rApoE4 was expressed in the E. Coli BL21(D3) strain and a soluble form of the protein was purified by a combination of affinity and size-exclusion chromatography that precluded a denaturation step. The structural integrity and the biochemical activity of the purified rApoE4 were confirmed by circular dichroism and a lipid-binding assay. Several biological parameters affected by rApoE4, such as mitochondrial morphology, mitochondrial membrane potential and reactive oxygen species production were studied in CNh cells, a neuronal cell line, and neurodifferentiation and dendritogenesis were analyzed in the SH-SY5Y neuroblastoma cell line. The improved rApoE4 purification technique reported here enables the production of highly purified protein that retain the structural properties and functional activity of the native protein, as confirmed by tests in two different neuronal cell lines in culture. 2023-06-28T16:00:41Z 2023-06-28T16:00:41Z 2023 Artículo Serrano, E., Barrantes, F.J., Valdivieso, A.G. Apolipoprotein E4 heterologous expression, purification under non-denaturing conditions, and effects on neuronal clonal cell lines [en línea]. Protein Expression and Purification. 2023, 210, doi:10.1016/j.pep.2023.106312 Disponible en: https://repositorio.uca.edu.ar/handle/123456789/16628 1046-5928 https://repositorio.uca.edu.ar/handle/123456789/16628 10.1016/j.pep.2023.106312 37236517 spa Acceso Restringido http://creativecommons.org/licenses/by-nc-nd/4.0/ application/pdf Elsevier Protein Expression and Purification. 2023, 210
institution Universidad Católica Argentina
institution_str I-33
repository_str R-139
collection Repositorio Institucional de la Universidad Católica Argentina (UCA)
language Español
topic ENFERMEDAD DE ALZHEIMER
PROTEINAS
APOLIPOPROTEINA
spellingShingle ENFERMEDAD DE ALZHEIMER
PROTEINAS
APOLIPOPROTEINA
Serrano, Ezequiel
Barrantes, Francisco José
Valdivieso, Ángel Gabriel
Apolipoprotein E4 heterologous expression, purification under non-denaturing conditions, and effects on neuronal clonal cell lines
topic_facet ENFERMEDAD DE ALZHEIMER
PROTEINAS
APOLIPOPROTEINA
description Abstract: The ε4 allele of the apolipoprotein E gene (APOE4) constitutes the main genetic risk factor for late-onset Alzheimer disease (AD). High amounts of pure apolipoprotein E4 (ApoE4), in a rapid and reproducible fashion, could be of value for studying its pathophysiological roles in AD. The aim of the present work was to optimize a preparative method to obtain highly purified recombinant ApoE4 (rApoE4) with full biological activity. rApoE4 was expressed in the E. Coli BL21(D3) strain and a soluble form of the protein was purified by a combination of affinity and size-exclusion chromatography that precluded a denaturation step. The structural integrity and the biochemical activity of the purified rApoE4 were confirmed by circular dichroism and a lipid-binding assay. Several biological parameters affected by rApoE4, such as mitochondrial morphology, mitochondrial membrane potential and reactive oxygen species production were studied in CNh cells, a neuronal cell line, and neurodifferentiation and dendritogenesis were analyzed in the SH-SY5Y neuroblastoma cell line. The improved rApoE4 purification technique reported here enables the production of highly purified protein that retain the structural properties and functional activity of the native protein, as confirmed by tests in two different neuronal cell lines in culture.
format Artículo
author Serrano, Ezequiel
Barrantes, Francisco José
Valdivieso, Ángel Gabriel
author_facet Serrano, Ezequiel
Barrantes, Francisco José
Valdivieso, Ángel Gabriel
author_sort Serrano, Ezequiel
title Apolipoprotein E4 heterologous expression, purification under non-denaturing conditions, and effects on neuronal clonal cell lines
title_short Apolipoprotein E4 heterologous expression, purification under non-denaturing conditions, and effects on neuronal clonal cell lines
title_full Apolipoprotein E4 heterologous expression, purification under non-denaturing conditions, and effects on neuronal clonal cell lines
title_fullStr Apolipoprotein E4 heterologous expression, purification under non-denaturing conditions, and effects on neuronal clonal cell lines
title_full_unstemmed Apolipoprotein E4 heterologous expression, purification under non-denaturing conditions, and effects on neuronal clonal cell lines
title_sort apolipoprotein e4 heterologous expression, purification under non-denaturing conditions, and effects on neuronal clonal cell lines
publisher Elsevier
publishDate 2023
url https://repositorio.uca.edu.ar/handle/123456789/16628
work_keys_str_mv AT serranoezequiel apolipoproteine4heterologousexpressionpurificationundernondenaturingconditionsandeffectsonneuronalclonalcelllines
AT barrantesfranciscojose apolipoproteine4heterologousexpressionpurificationundernondenaturingconditionsandeffectsonneuronalclonalcelllines
AT valdiviesoangelgabriel apolipoproteine4heterologousexpressionpurificationundernondenaturingconditionsandeffectsonneuronalclonalcelllines
_version_ 1770442213336023040

Ejemplares similares