Expression,purification and functional characterization of the two zinc-fingers domain of the human GATA1

I APEZTEGUIA; R CALLIGARIS; S BOTTARDI; Claudio Ventura SANTORO

doi:10.1006/prep.1994.1074

Expression,purification and functional characterization of the two zinc-fingers domain of the human GATA1

I APEZTEGUIA
, R CALLIGARIS
, S BOTTARDI
, Claudio Ventura SANTORO

Department of Health Sciences

Research output: Contribution to journal › Article › peer-review

Abstract

The DNA-binding domain of the erythroid transcription factor GATA-1 consists of two closely related, but distinct zinc-fingers which are highly conserved among the members of the growing family of GATA-like factors. The DNA-binding domain of the human GATA-1 (F₁F₂) was expressed as a histidine-tagged fusion protein in Escherichia coli. The denaturated protein was purified by Ni²⁺-chelate affinity chromatography and renaturated in situ. The active recombinant protein was purified by DNA affinity chromatography. F₁F₂ displayed GATA-1 specific binding activity toward its DNA recognition sequences within the hypersensitive site 3 of the human locus control region and the human γ-globin promoter. In contrast to GATA-1 protein purified from K562 nuclei, the recombinant F₁F₂ bound also the CCAAT-box region of the human γ-globin promoter.

Original language	English
Pages (from-to)	101-107
Number of pages	7
Journal	Protein Expression and Purification
Volume	5
DOIs	https://doi.org/10.1006/prep.1994.1074
Publication status	Published - 1994

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10.1006/prep.1994.1074

http://hdl.handle.net/11579/4826

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title = "Expression,purification and functional characterization of the two zinc-fingers domain of the human GATA1",

abstract = "The DNA-binding domain of the erythroid transcription factor GATA-1 consists of two closely related, but distinct zinc-fingers which are highly conserved among the members of the growing family of GATA-like factors. The DNA-binding domain of the human GATA-1 (F1F2) was expressed as a histidine-tagged fusion protein in Escherichia coli. The denaturated protein was purified by Ni2+-chelate affinity chromatography and renaturated in situ. The active recombinant protein was purified by DNA affinity chromatography. F1F2 displayed GATA-1 specific binding activity toward its DNA recognition sequences within the hypersensitive site 3 of the human locus control region and the human γ-globin promoter. In contrast to GATA-1 protein purified from K562 nuclei, the recombinant F1F2 bound also the CCAAT-box region of the human γ-globin promoter.",

author = "I APEZTEGUIA and R CALLIGARIS and S BOTTARDI and SANTORO, \{Claudio Ventura\}",

year = "1994",

doi = "10.1006/prep.1994.1074",

language = "English",

volume = "5",

pages = "101--107",

journal = "Protein Expression and Purification",

issn = "1046-5928",

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T1 - Expression,purification and functional characterization of the two zinc-fingers domain of the human GATA1

AU - APEZTEGUIA, I

AU - CALLIGARIS, R

AU - BOTTARDI, S

AU - SANTORO, Claudio Ventura

PY - 1994

Y1 - 1994

N2 - The DNA-binding domain of the erythroid transcription factor GATA-1 consists of two closely related, but distinct zinc-fingers which are highly conserved among the members of the growing family of GATA-like factors. The DNA-binding domain of the human GATA-1 (F1F2) was expressed as a histidine-tagged fusion protein in Escherichia coli. The denaturated protein was purified by Ni2+-chelate affinity chromatography and renaturated in situ. The active recombinant protein was purified by DNA affinity chromatography. F1F2 displayed GATA-1 specific binding activity toward its DNA recognition sequences within the hypersensitive site 3 of the human locus control region and the human γ-globin promoter. In contrast to GATA-1 protein purified from K562 nuclei, the recombinant F1F2 bound also the CCAAT-box region of the human γ-globin promoter.

AB - The DNA-binding domain of the erythroid transcription factor GATA-1 consists of two closely related, but distinct zinc-fingers which are highly conserved among the members of the growing family of GATA-like factors. The DNA-binding domain of the human GATA-1 (F1F2) was expressed as a histidine-tagged fusion protein in Escherichia coli. The denaturated protein was purified by Ni2+-chelate affinity chromatography and renaturated in situ. The active recombinant protein was purified by DNA affinity chromatography. F1F2 displayed GATA-1 specific binding activity toward its DNA recognition sequences within the hypersensitive site 3 of the human locus control region and the human γ-globin promoter. In contrast to GATA-1 protein purified from K562 nuclei, the recombinant F1F2 bound also the CCAAT-box region of the human γ-globin promoter.

UR - https://iris.uniupo.it/handle/11579/4826

U2 - 10.1006/prep.1994.1074

DO - 10.1006/prep.1994.1074

M3 - Article

SN - 1046-5928

VL - 5

SP - 101

EP - 107

JO - Protein Expression and Purification

JF - Protein Expression and Purification

ER -

Expression,purification and functional characterization of the two zinc-fingers domain of the human GATA1

Abstract

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