Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Experiment Videos

Evidence for an intermediate with a single-strand break in the reaction catalyzed by the DNA untwisting enzyme.

J J Champoux

    Proceedings of the National Academy of Sciences of the United States of America
    |October 1, 1976
    PubMed
    Summary
    This summary is machine-generated.

    Related Concept Videos

    You might also read

    Related Articles

    Articles linked to this work by shared authors, journal, and citation graph.

    Sort by
    Same author

    Purification of baculovirus-expressed human DNA topoisomerase I.

    Methods in molecular biology (Clifton, N.J.)·2003
    Same author

    The tyrosyl-DNA phosphodiesterase Tdp1 is a member of the phospholipase D superfamily.

    Proceedings of the National Academy of Sciences of the United States of America·2001
    Same author

    DNA topoisomerases: structure, function, and mechanism.

    Annual review of biochemistry·2001
    Same author

    Structural alterations in the DNA ahead of the primer terminus during displacement synthesis by reverse transcriptases.

    Journal of molecular biology·2001
    Same author

    Structure-based analysis of the effects of camptothecin on the activities of human topoisomerase I.

    Annals of the New York Academy of Sciences·2001
    Same author

    Assaying DNA topoisomerase I relaxation activity.

    Methods in molecular biology (Clifton, N.J.)·2000

    This study identifies a nicked DNA intermediate in the DNA untwisting enzyme

    Area of Science:

    • Biochemistry
    • Molecular Biology
    • Enzymology

    Background:

    • DNA topology is crucial for cellular processes.
    • DNA untwisting enzymes (topoisomerases) regulate DNA topology.
    • Understanding enzyme mechanisms requires identifying reaction intermediates.

    Purpose of the Study:

    • To detect and characterize the nicked DNA intermediate in the DNA untwisting reaction.
    • To provide evidence supporting the sequential mechanism of DNA relaxation by the enzyme.

    Main Methods:

    • Utilized highly purified rat liver nuclear enzyme.
    • Assessed nicking of simian virus 40 DNA using alkaline sucrose gradient sedimentation and CsCl-propidium diiodide equilibrium centrifugation.
    • Manipulated salt concentrations (KCl) to study enzyme activity and intermediate stability.

    Related Experiment Videos

    Main Results:

    • Successfully detected a nicked DNA intermediate during the untwisting reaction.
    • Nicking occurred within the active salt concentration range and ceased at higher salt concentrations.
    • The nicked intermediate was transient, disappearing upon increasing salt concentration.
    • Nicking was independent of DNA concentration, and unlabeled DNA could chase labeled nicked DNA.

    Conclusions:

    • The observed nicked DNA represents a genuine intermediate in the DNA untwisting enzyme's reaction mechanism.
    • These findings support a sequential strand-breaking and rejoining model for DNA relaxation.