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Related Concept Videos

Lagging Strand Synthesis01:59

Lagging Strand Synthesis

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During replication, the complementary strands in double-stranded DNA are synthesized at different rates. Replication first begins on the leading strand. Replication starts later, occurs more slowly, and proceeds discontinuously on the lagging strand.
There are several major differences between synthesis of the leading strand and synthesis of the lagging strand. 1) Leading strand synthesis happens in the direction of replication fork opening, whereas lagging strand synthesis happens in the...
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The Replisome03:01

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DNA replication is carried out by a large complex of proteins that act in a coordinated matter to achieve high-fidelity DNA replication. Together this complex is known as the DNA replication machinery or the replisome.
The synthesis of the leading and lagging strands is a highly coordinated process. To explain this, the “Trombone model” was proposed by Bruce Alberts in 1980. The DNA loop formation starts when a primer is synthesized on the parent lagging strand. The loop grows with...
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DNA Replication02:40

DNA Replication

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DNA replication involves the separation of the two strands of the double helix, with each strand serving as a template from which the new complementary strand is copied.  After replication, each double-stranded DNA includes one parental or “old” strand and one “new” strand. This is known as semiconservative replication. The resulting DNA molecules have the same sequence and are divided equally into the two daughter cells.
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Restarting Stalled Replication Forks02:37

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DNA replication is initiated at sites containing predefined DNA sequences known as origins of replication. DNA is unwound at these sites by the minichromosome maintenance (MCM) helicase and other factors such as Cdc45 and the associated GINS complex.The unwound single strands are protected by replication protein A (RPA) until DNA polymerase starts synthesizing DNA at the 5’ end of the strand in the same direction as the replication fork. To prevent the replication fork from falling apart,...
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Proofreading01:31

Proofreading

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Synthesis of new DNA molecules is carried out by the enzyme DNA polymerase, which adds nucleotides on the daughter strand complementary to the template DNA strand. DNA polymerase has a higher affinity to add the correct base and ensures fidelity during DNA replication. Furthermore,  it exhibits proofreading activity during replication, using an exonuclease domain that cuts off incorrect nucleotides from the nascent DNA strand.
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Translesion DNA Polymerases02:10

Translesion DNA Polymerases

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Translesion (TLS) polymerases rescue stalled DNA polymerases at sites of damaged bases by replacing the replicative polymerase and installing a nucleotide across the damaged site. Doing so, TLS allows additional time for the cell to repair the damage before resuming regular DNA replication.
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Related Experiment Video

Updated: Sep 17, 2025

Single-Molecule Fluorescence Visualization of DNA Polymerase Dynamics at G-Quadruplexes
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Single-Molecule Fluorescence Visualization of DNA Polymerase Dynamics at G-Quadruplexes

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Visualizing DNA polymerase ι catalyze Hoogsteen-directed DNA synthesis.

Zach Frevert1, Devin T Reusch1, Melissa S Gildenberg1

  • 1Department of Biochemistry and Molecular Biology, University of Iowa College of Medicine, Iowa City, IA, USA.

Nature Communications
|July 2, 2025
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Summary

Translesion synthesis polymerase ι (Pol ι) uses Hoogsteen base pairs for nucleotide incorporation opposite DNA lesions. This enzyme also exhibits pyrophosphatase activity, aiding in DNA synthesis and translocation.

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Structural Biology

Background:

  • Translesion synthesis (TLS) polymerases are crucial for DNA repair, bypassing damaged DNA segments.
  • Pol ι is a TLS polymerase known to incorporate nucleotides opposite various DNA adducts.
  • Previous studies revealed Pol ι's use of Hoogsteen base pairs during nucleotide binding but lacked mechanistic detail on incorporation.

Purpose of the Study:

  • To elucidate the detailed mechanism of nucleotide incorporation catalyzed by Pol ι.
  • To visualize the dynamic process of phosphodiester bond formation during Pol ι activity.
  • To identify reaction intermediates and understand the role of Pol ι's active site in DNA synthesis.

Main Methods:

  • Time-lapse X-ray crystallography to capture dynamic structural changes.
  • Molecular dynamics simulations to model enzyme-nucleotide interactions.
  • Biochemical assays to characterize enzyme activity.

Main Results:

  • Pol ι maintains Hoogsteen base pairing throughout the nucleotide incorporation process.
  • The enzyme exhibits intrinsic pyrophosphatase activity, cleaving dNTPs into monophosphates within the active site.
  • Structural and dynamic analyses revealed key active site features facilitating DNA translocation and processive synthesis.

Conclusions:

  • Pol ι employs a unique Hoogsteen base-pairing strategy for efficient and accurate nucleotide incorporation opposite DNA lesions.
  • The pyrophosphatase activity of Pol ι contributes to its catalytic mechanism and processivity.
  • Understanding Pol ι's mechanism provides insights into DNA repair pathways and potential therapeutic targets.