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Giới thiệu nội dung

Purification And Functional Characterization Of Human 11ẞ Hydroxylase Expressed In Escherichia Coli

Tác giả: Andy Zöllner, Norio Kagawa, Michael R. Waterman, Yasuki Nonaka, Koji Takio, Yoshitsugu Shiro, Frank Hannemann, and Rita Bernhardt

Lĩnh vực: Biochemistry

Nội dung tài liệu: This study reports on the successful and reproducible purification of recombinant human 11β-hydroxylase (hCYP11B1) expressed in Escherichia coli, coexpressed with molecular chaperones GroES/GroEL. The purified protein was characterized for its functionality using various methods, including substrate conversion assays, stopped-flow experiments, and Biacore measurements. The research demonstrates that the enzyme effectively hydroxylates its substrates at the 11-beta position and provides insights into its interaction with its redox partner, adrenodoxin. This work establishes a foundation for further investigation into the structure and function of this physiologically important enzyme.

Mục lục chi tiết:

  • Keywords
  • Correspondence
  • Abbreviations
  • Abstract
  • Introduction
  • Results and Discussion
  • Experimental procedures
    • Protein expression and purification
    • RP-HPLC and mass spectrometry of the purified hCYP11B1
    • UV/visible and CD spectroscopy
    • Enzyme activity assays
    • Optical biosensor measurements
    • Kinetics by rapid mixing
  • Acknowledgements
  • References