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Related Experiment Videos

Methods to Screen for Radical SAM Enzyme Crystallization Conditions.

Lydie Martin1, Xavier Vernède1, Yvain Nicolet2

  • 1Univ. Grenoble Alpes, CEA, CNRS, IBS, Metalloproteins Unit, Grenoble, France.

Methods in Molecular Biology (Clifton, N.J.)
|July 22, 2021
PubMed
Summary

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Nature chemistry·2024

Radical S-adenosyl-L-methionine (SAM) enzymes are crucial metalloenzymes. New protocols enable rapid screening of crystallization conditions for these challenging proteins, aiding biotechnological applications.

Area of Science:

  • Biochemistry
  • Structural Biology
  • Enzymology

Background:

  • Radical S-adenosyl-L-methionine (SAM) proteins represent a vast superfamily of metalloenzymes.
  • These enzymes catalyze complex biochemical reactions and are vital in natural product biosynthesis.
  • Structural elucidation of radical SAM enzymes is often hindered by crystallization difficulties.

Purpose of the Study:

  • To present novel protocols and equipment for rapid screening of crystallization conditions for radical SAM enzymes.
  • To demonstrate the utility of these methods under anaerobic conditions.
  • To highlight the application of these techniques in the structural study of the nitrogenase maturase NifB.

Main Methods:

  • Development of specialized equipment for high-throughput screening.
Keywords:
Anaerobic crystallizationCrystal flash cooling. FeMo coassemblyFe-S cluster reconstitutionFeMo cofactorMetalloproteinProtein expression and purification

Related Experiment Videos

  • Implementation of anaerobic conditions to maintain enzyme activity and facilitate crystallization.
  • Application of screening protocols to the nitrogenase maturase NifB.

    • Main Results:

    • Successful development of protocols for rapid crystallization screening of radical SAM enzymes.
    • Demonstration of effective screening under strictly anaerobic conditions.
    • Obtained crystallization conditions for the nitrogenase maturase NifB, facilitating structural studies.

    Conclusions:

    • The developed protocols and equipment significantly improve the efficiency of crystallizing challenging radical SAM enzymes.
    • Anaerobic screening is crucial for studying oxygen-sensitive metalloenzymes like NifB.
    • These advancements hold promise for broader biotechnological applications of radical SAM enzymes.