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富玛莱酸氢酶含含蛋白质的表达、纯化、结晶和酶测定
JoVE Journal
Bioquímica
This content is Free Access.
JoVE Journal Bioquímica
Expression, Purification, Crystallization, and Enzyme Assays of Fumarylacetoacetate Hydrolase Domain-Containing Proteins
DOI:

10:21 min

June 20, 2019

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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

Summary

Tadução automática

用实例描述了含紫酯水酶的表达和纯化(大肠杆菌、FPLC中的表达)。纯化蛋白用于结晶和抗体生产,并用于酶测定。提出了选定的光度测定法,以显示FAHD1的多功能性,如牛酸脱碳酶和丙烯酰胺水酶。

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