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Ekspression, rensning, krystallisering og enzym assays af Fumarylacetoacetat hydrolase-domæne-holdige proteiner
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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
Capítulos
- 00:04Título
- 01:12Expression of FAHD Proteins in Competent E. coli
- 03:44Lysis of Bacterial Pellets and Filtration of Debris
- 04:33Purification of His-tagged FAHD Proteins using Ni-NTA Affinity Chromatography
- 05:59Purification of FAHD Proteins via Ion Exchange Chromatography
- 06:35Basic FAHD Activity Assays with Substrates Oxaloacetate and Acetylpyruvate
- 07:53Results: Expression and Purification of Fumarylacetoacetate Hydrolase Domain-containing Proteins
- 09:19Conclusion
Udtryk og rensning af fumarylacetoacetat lav affinitet domæne-holdige proteiner er beskrevet med eksempler (udtryk i E. coli, fplc). Rensede proteiner bruges til krystallisering og antistof produktion og anvendes til enzym assays. Udvalgte fotometriske analyser er præsenteret for at vise multi-funktionaliteten af FAHD1 som oxaloacetat decarboxylase og acylpyruvate hydrolase.
