Related Experiment Video

Updated: May 1, 2026

Atomic Force Microscopy Investigations of DNA Lesion Recognition in Nucleotide Excision Repair
10:59

Atomic Force Microscopy Investigations of DNA Lesion Recognition in Nucleotide Excision Repair

Published on: May 24, 2017

9.3K

The Mus81-Mms4 structure-selective endonuclease requires nicked DNA junctions to undergo conformational changes and

Sucheta Mukherjee1, William Douglass Wright1, Kirk Tevebaugh Ehmsen1

  • 1Department of Microbiology & Molecular Genetics, University of California, One Shields Ave., Davis, Davis CA 95616-8665, USA.

Nucleic Acids Research
|April 19, 2014
PubMed

Insights

Budding yeast Mus81-Mms4/EME1, a DNA endonuclease, requires flexible DNA junctions for cleavage. It induces a sharp bend in DNA, facilitating precise cutting during homologous recombination.

Area of Science:

  • Molecular Biology
  • Biochemistry
  • Genetics

Background:

  • Mus81-Mms4/EME1 is a crucial endonuclease involved in cleaving joint DNA molecules during homologous recombination in both mitotic and meiotic cells.
  • Understanding the catalytic mechanism of Mus81-Mms4/EME1 is essential for comprehending DNA repair and genome stability.

Purpose of the Study:

  • To investigate the kinetic and structural requirements for Mus81-Mms4/EME1 DNA cleavage activity.
  • To elucidate the mechanism by which Mus81-Mms4/EME1 recognizes and processes specific DNA structures.

Main Methods:

  • Kinetic analysis using physically tethered DNA substrates.
  • Förster Resonance Energy Transfer (FRET) experiments to study DNA-protein interactions.
  • Thiol crosslinking to detect conformational changes in Mus81-Mms4/EME1 upon DNA binding.

Main Results:

  • Mus81-Mms4/EME1 requires inherent rotational flexibility in DNA junctions for optimal catalysis.
  • Substrate recognition involves the induction of a sharp bend (100°) in DNA by Mus81-Mms4/EME1.
  • DNA binding induces a conformational change in the Mus81-Mms4/EME1 heterodimer, suggesting a substrate-induced conformational transition.

Conclusions:

  • A model is proposed where Mus81-Mms4/EME1 binding is followed by a rate-limiting conformational change in both protein and substrate.
  • This interaction leads to a kinked DNA molecule, positioning the nuclease for cleavage between the fourth and fifth nucleotide.
  • Mutually compatible conformational changes between Mus81-Mms4/EME1 and its substrates optimize incision activity on nicked junction molecules.

More Related Videos

Author Spotlight: Characterizing Novel Enzymes from Extremophiles and Common Pathogens to Understand DNA Repair and Replication05:33

Author Spotlight: Characterizing Novel Enzymes from Extremophiles and Common Pathogens to Understand DNA Repair and Replication

Published on: July 5, 2024

1.7K
Single-Molecule Dwell-Time Analysis of Restriction Endonuclease-Mediated DNA Cleavage09:53

Single-Molecule Dwell-Time Analysis of Restriction Endonuclease-Mediated DNA Cleavage

Published on: February 7, 2021

1.8K

Related Concept Videos

Single-Strand DNA Binding Proteins01:03

Single-Strand DNA Binding Proteins

For successful DNA replication, the unwinding of double-stranded DNA must be accompanied by stabilization and protection of the separated single strands of the DNA. This crucial task is performed by single-strand DNA-binding (SSB) proteins. They bind to the DNA in a sequence-independent manner, which means that the nitrogenous bases of the DNA need not be present in a specific order for binding of SSB proteins to it. The binding of SSB proteins straightens single-stranded DNA (ssDNA) and makes...
12.9K
Restriction Enzymes01:11

Restriction Enzymes

Restriction enzymes are bacterial enzymes used to cut DNA in a sequence-specific manner. To cleave DNA, they bind to specific palindromic sequences called restriction sites. Such palindromic DNA sequences or inverted repeats are commonly found in regions of functional significance, such as the origin of replication, gene operator sites, and regions containing transcription termination signals.
The host bacteria protect their own genomic DNA from these enzymes by methylating these sites. Some...
31.4K
Nucleotide Excision Repair01:08

Nucleotide Excision Repair

Overview
33.7K
Nucleotide Excision Repair01:38

Nucleotide Excision Repair

DNA Distortion and Damage
Cells are regularly exposed to mutagens—factors in the environment that can damage DNA and generate mutations. UV radiation is one of the most common mutagens and is estimated to introduce a significant number of changes in DNA. These include bends or kinks in the structure, which can block DNA replication or transcription. If these errors are not fixed, the damage can cause mutations, which in turn can result in cancer or disease depending on which sequences are...
4.6K
Homologous Recombination02:31

Homologous Recombination

The basic reaction of homologous recombination (HR) involves two chromatids that contain DNA sequences sharing a significant stretch of identity. One of these sequences uses a strand from another as a template to synthesize DNA in an enzyme-catalyzed reaction. The final product is a novel amalgamation of the two substrates. To ensure an accurate recombination of sequences, HR is restricted to the S and G2 phases of the cell cycle. At these stages, the DNA has been replicated already and the...
58.8K
Mismatch Repair01:20

Mismatch Repair

Organisms are capable of detecting and fixing nucleotide mismatches that occur during DNA replication. This sophisticated process requires identifying the new strand and replacing the erroneous bases with correct nucleotides. Mismatch repair is coordinated by many proteins in both prokaryotes and eukaryotes.
The Mutator Protein Family Plays a Key Role in DNA Mismatch Repair
The human genome has more than 3 billion base pairs of DNA per cell. Prior to cell division, that vast amount of genetic...
5.4K