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

In eukaryotic flap endonuclease 1, the C terminus is essential for substrate binding.

M Stucki1, Z O Jónsson, U Hübscher

  • 1Institut für Veterinärbiochemie, Universität Zürich, Winterthurerstrasse 190, CH-8057 Zürich, Switzerland.

The Journal of Biological Chemistry
|November 18, 2000
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

[Is health really the most important value? - Results of a representative survey of the German general population concerning the subjective meaning of health].

Gesundheitswesen (Bundesverband der Arzte des Offentlichen Gesundheitsdienstes (Germany))·2010
Same author

DNA polymerases and oxidative damage: friends or foes?

Current molecular pharmacology·2009
Same author

Investigations of prion and virus safety of a new liquid IVIG product.

Biologicals : journal of the International Association of Biological Standardization·2008
Same author

DNA damage response mediators MDC1 and 53BP1: constitutive activation and aberrant loss in breast and lung cancer, but not in testicular germ cell tumours.

Oncogene·2007
Same author

Modulation of cellular response to cisplatin by a novel inhibitor of DNA polymerase beta.

Molecular pharmacology·2005
Same author

Reconstitution of the base excision repair pathway for 7,8-dihydro-8-oxoguanine with purified human proteins.

Nucleic acids research·2002

Flap endonuclease 1 (Fen1) is crucial for DNA replication and repair. Deleting its C-terminal basic region impairs enzymatic activity and substrate binding without affecting PCNA interaction, revealing a key regulatory domain.

Area of Science:

  • Molecular Biology
  • Biochemistry
  • Genetics

Background:

  • Flap endonuclease 1 (Fen1) is a structure-specific metallonuclease vital for DNA replication and repair.
  • Fen1 interacts with proliferating cell nuclear antigen (PCNA), which stimulates its enzymatic activity in vitro.
  • The PCNA interaction site on Fen1 is near its C terminus, adjacent to a conserved basic region in eukaryotes.

Purpose of the Study:

  • To investigate the functional roles of the C-terminal PCNA interaction motif and adjacent basic region in human Fen1.
  • To analyze the impact of deleting these regions on Fen1's enzymatic activity, PCNA interaction, and substrate binding.

Main Methods:

  • Construction and analysis of two human Fen1 deletion mutants.
  • Enzymatic activity assays.

Related Experiment Videos

  • Electrophoretic mobility shift assays (EMSA) to assess substrate binding.
  • Analysis of protein-PCNA interactions.

    • Main Results:

    • Deletion of the C-terminal basic region did not abolish PCNA interaction.
    • This deletion mutant exhibited significantly reduced enzymatic activity.
    • EMSA revealed a severe defect in substrate binding for the C-terminal deletion mutant.
    • The PCNA interaction motif deletion mutant was not explicitly detailed in its results, but the focus was on the C-terminal region.

    Conclusions:

    • The C terminus of eukaryotic Fen1 contains at least two functionally distinct regions.
    • These regions, including the basic C-terminal region, play crucial roles in regulating Fen1's enzymatic activity and substrate binding.
    • These findings suggest a complex regulatory mechanism involving the Fen1 C terminus for DNA metabolic processes.