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Related Experiment Videos

An engineered chorismate mutase with allosteric regulation.

Sheng Zhang1, David B Wilson, Bruce Ganem

  • 1Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY 14853, USA.

Bioorganic & Medicinal Chemistry
|June 24, 2003
PubMed
Summary

Researchers engineered a bifunctional P-protein from E. coli, removing the prephenate dehydratase domain. This created a functional chorismate mutase linked to a phenylalanine binding domain, allowing study of its catalytic properties.

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Area of Science:

  • Biochemistry
  • Enzyme Engineering
  • Protein Catalysis

Background:

  • The bifunctional P-protein from E. coli is crucial for phenylalanine biosynthesis.
  • It comprises three distinct domains: chorismate mutase, prephenate dehydratase, and an allosteric feedback inhibition domain.
  • These domains retain function when separated, making P-protein a model for engineering allosteric regulation.

Purpose of the Study:

  • To investigate the engineering of allosteric effects into protein catalysts using modular regulatory elements.
  • To characterize the catalytic properties of a modified P-protein lacking the prephenate dehydratase domain.
  • To assess the impact of phenylalanine on the engineered chorismate mutase activity.

Main Methods:

  • Deletion of the prephenate dehydratase domain from the bifunctional P-protein.

Related Experiment Videos

  • Engineering and overexpression of a functional chorismate mutase linked to the phenylalanine binding domain.
  • Analysis of the catalytic properties of the engineered enzyme in the presence and absence of phenylalanine.

    • Main Results:

    • A modified P-protein was successfully engineered, retaining chorismate mutase activity.
    • The catalytic properties of the isolated chorismate mutase domain were characterized.
    • The influence of phenylalanine on the enzyme's activity was investigated.

    Conclusions:

    • The study demonstrates the feasibility of engineering allosteric regulation into enzymes by combining functional domains.
    • The engineered chorismate mutase provides a simplified system for studying enzyme kinetics and allosteric inhibition.
    • This work contributes to understanding protein engineering principles for creating novel biocatalysts.