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Journal / Biochemistry / Expression, Purification, Crystallization, and…
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Biochemistry

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JoVE Journal Biochemistry
Expression, Purification, Crystallization, and Enzyme Assays of Fumarylacetoacetate Hydrolase Domain-Containing Proteins
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Expression, Purification, Crystallization, and Enzyme Assays of Fumarylacetoacetate Hydrolase Domain-Containing Proteins

Article DOI: 10.3791/59729-v • 10:21 min • June 20th, 2019
June 20th, 2019 •
1,2, 1,2, 1,2, 1, 1, 3, 3, 4, 1,2
1Research Institute for Biomedical Aging Research, University of Innsbruck Austria, 2Center for Molecular Biosciences Innsbruck (CMBI), University of Innsbruck Austria, 3Division of Genetic Epidemiology, Medical University of Innsbruck Austria, 4Faculty of Chemistry, Department of Organic Chemistry, University of Vienna Austria

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Summary
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Expression and purification of fumarylacetoacetate hydrolase domain-containing proteins is described with examples (expression in E. coli, FPLC). Purified proteins are used for crystallization and antibody production and employed for enzyme assays. Selected photometric assays are presented to display the multi-functionality of FAHD1 as oxaloacetate decarboxylase and acylpyruvate hydrolase.

Tags

Expression Purification Crystallization Enzyme Assays Fumarylacetoacetate Hydrolase Domain-containing Proteins FAH Domain Protein Structure Molecular Mechanisms Cellular Senescence IPTG Inducible Vector Systems Poly-histidine Type Proteins Nickel NTA Agarose Resins Affinity Chromatography FPLC Plasmid BL21-DE3 PLysS E.coli Bacteria
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