Name: proofreading in dna replication
Uploaded: 2020-11-23T11:47:35.000Z
Duration: 1 min 43 s
Channel: JoVE

K12 Standards

Alabama Course of Study Science

9-12

Biology

from-molecules-to-organisms-structures-and-processes

From Molecules to Organisms: Structures and Processes

9-12.B.1.3

Proofreading

During DNA replication, free nucleotides in the cell are added to the new strand in a sequence complementary to the template. Adenine pairs with thymine, and cytosine pairs with guanine.
 
 Sometimes, nucleotides pair incorrectly, such as when adenine pairs with cytosine. DNA polymerase proofreading helps fix these errors and reduces the chance of mutations. If they are not corrected, these errors can cause disease.
 
 First, DNA polymerase binds more strongly to correctly paired nucleotides. This reduces the chance of incorrect pairings.
 
 Second, as nucleotides begin pairing with the template, DNA polymerase undergoes a conformational change. This change causes incorrectly paired nucleotides to detach, allowing the correct nucleotides to be added.
 
 Third, if an incorrect nucleotide is added to the three-prime end of the growing DNA chain, the mismatch distorts the DNA structure and pauses DNA polymerase movement.
 
 DNA polymerase has three-prime-to-five-prime exonuclease activity, which lets it remove an incorrectly added nucleotide. It then inserts the correct nucleotide. This process is called exonucleolytic proofreading.
 
 These steps help keep DNA replication accurate by separating nucleotide selection from error correction.

Barbari, Stephanie R. et Polina V. Shcherbakova. « Défauts de l’ADN polymérase répliquative dans les cancers humains : conséquences, mécanismes et implications pour la thérapie. » Réparation de l’ADN 56 (2017) : 16-25. [<a href="https://doi.org/10.1016/j.dnarep.2017.06.003">Source</a>]

relecture

<h4>Aper&#xE7;u</h4>
 
 La synth&#xE8;se de nouvelles mol&#xE9;cules d&#x2019;ADN commence lorsque la polym&#xE9;rase d&#x2019;ADN relie les nucl&#xE9;otides dans une s&#xE9;quence compl&#xE9;mentaire au brin d&#x2019;ADN matrice. L&#x2019;ADN polym&#xE9;rase a plus d&#x2019;affinit&#xE9; pour la base correcte afin d&#x2019;assurer la fid&#xE9;lit&#xE9; lors de la r&#xE9;plication de l&#x2019;ADN. L&#x2019;ADN polym&#xE9;rase fait en outre la relecture pendant la r&#xE9;plication, &#xE0; l&#x2019;aide d&#x2019;un domaine d&#x2019;exonucl&#xE9;ase qui coupe les nucl&#xE9;otides incorrects du brin d&#x2019;ADN naissant.
 
 <h4>Les erreurs pendant la r&#xE9;plication sont corrig&#xE9;es par l&#x2019;enzyme d&#x2019;ADN polym&#xE9;rase</h4>
 
 L&#x2019;ADN g&#xE9;nomique est synth&#xE9;tis&#xE9; dans le sens de 5&#x2019; &#xE0; 3&#x2019;. Chaque cellule contient un certain nombre de polym&#xE9;rases d&#x2019;ADN qui jouent des r&#xF4;les diff&#xE9;rents dans la synth&#xE8;se et la correction des erreurs dans l&#x2019;ADN ; l&#x2019;ADN polym&#xE9;rase delta et epsilon poss&#xE8;dent la facult&#xE9; de relecture au moment de la r&#xE9;plication de l&#x2019;ADN nucl&#xE9;aire. Ces polym&#xE9;rases &#x201C; lisent &#x201D; chaque base apr&#xE8;s qu&#x2019;elle soit ajout&#xE9;e au nouveau brin. Si la base nouvellement ajout&#xE9;e est incorrecte, la polym&#xE9;rase inverse la direction (passant de 3&#x2019; &#xE0; 5&#x2019;)&#xA0;et utilise un domaine exonucl&#xE9;olytique pour couper la base incorrecte. Par la suite, elle est remplac&#xE9;e par la base correcte.
 
 <h4>Les mutations dans le domaine de l&#x2019;exonucl&#xE9;ase de l&#x2019;ADN polym&#xE9;rase sont li&#xE9;es aux cancers</h4>
 
 La relecture est importante pour emp&#xEA;cher que des mutations se produisent dans l&#x2019;ADN nouvellement synth&#xE9;tis&#xE9;, mais que se passe-t-il lorsque le m&#xE9;canisme de relecture &#xE9;choue ? Lorsqu&#x2019;une mutation modifie le domaine de l&#x2019;exonucl&#xE9;ase de l&#x2019;ADN polym&#xE9;rase, elle perd la facult&#xE9; d&#x2019;&#xE9;liminer les nucl&#xE9;otides incorrects. En cons&#xE9;quence, les mutations peuvent s&#x2019;accumuler rapidement dans tout le g&#xE9;nome. Ce type de mutation a &#xE9;t&#xE9; li&#xE9; &#xE0; divers types de cancer.
 
 <h4>La polym&#xE9;rase d&#x2019;ADN de faible fid&#xE9;lit&#xE9; peut g&#xE9;n&#xE9;rer des s&#xE9;quences d&#x2019;ADN mut&#xE9;es</h4>
 
 Des ADN polym&#xE9;rases modifi&#xE9;es sont utilis&#xE9;es en science de laboratoire pour la r&#xE9;action en cha&#xEE;ne par polym&#xE9;rase (PCR), une technique in vitro pour fabriquer de nombreuses copies de fragments sp&#xE9;cifiques d&#x2019;ADN. Alors que les polym&#xE9;rases de haute fid&#xE9;lit&#xE9; sont utilis&#xE9;es lorsqu&#x2019;il est important que le produit final soit parfait, certaines techniques, telles que la PCR sujettes aux erreurs, cherchent &#xE0; g&#xE9;n&#xE9;rer des mutations dans une portion d&#x2019;ADN expr&#xE8;s. Ces techniques utilisent des polym&#xE9;rases qui ont une facult&#xE9; de relecture affaiblie.

Relecture

Overview

Synthesis of new DNA molecules starts when DNA polymerase links nucleotides together in a sequence that is complementary to the template DNA ...

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Education

Enseignement

JoVE Core

Molecular Biology

Biologie moléculaire

Unit 1

Unité 1

DNA Replication

Réplication de l'ADN

KEY TERMS AND CONCEPTS

TERMES ET CONCEPTS CLÉS

proofreading in dna replication

Proofreading in DNA replication helps keep DNA copying accurate. It is a built-in check that reduces mistakes as new DNA is made.

DNA polymerase is the enzyme that carries out this checking during replication. When it adds a wrong nucleotide, it can remove the error and replace it with the correct one. This correction step helps maintain the correct DNA sequence.

Proofreading is an important part of DNA replication because it supports genetic accuracy. By catching mistakes as they happen, the cell lowers the chance that errors will remain in the finished DNA molecule.

Proofreading in DNA Replication

replication in prokaryotes

Prokaryotic DNA Replication Steps

Prokaryotic DNA replication copies the chromosome before a cell divides. In bacteria, this process starts at a specific DNA site called the origin of replication. From there, the cell builds new DNA strands using enzymes that match each base with its partner. The DNA double helix first opens so replication can begin. As the strands separate, each original strand serves as a template. New nucleotides are added in a set order, which helps the cell make an accurate copy of its genetic material. <br...

replication in eukaryotes

Eukaryotic DNA Replication Steps

Eukaryotic DNA replication copies the genome before a cell divides. In eukaryotes, this process happens in a controlled sequence so each new cell can receive genetic information. Replication begins at specific sites on the DNA called origins. The DNA is opened, and each original strand serves as a template, meaning it guides the building of a new complementary strand. This sets up two DNA molecules from one original molecule. The process then continues as enzymes add matching nucleotides to...

dna base pairing

DNA Base Pair Rules

DNA base pairing follows specific rules that help DNA carry genetic information. In DNA, the bases pair in a regular way, with adenine matching thymine and cytosine matching guanine. These pairs fit together because of their chemical shapes and bonding patterns. These base pairs also support accurate DNA replication. When DNA copies itself, each strand can guide the formation of a new partner strand using the pairing rules. Understanding base pairing helps explain how DNA stays organized and...

the dna replication fork

DNA Replication Stress and Fork Stalling

DNA replication stress and fork stalling can disrupt the copying of a cell’s genome. The replication fork is the Y-shaped site where DNA strands separate and new complementary strands are made at the same time. This linked unwinding and synthesis helps cells duplicate DNA efficiently and with fewer errors. In organisms with small circular DNA, such as E. coli, replication often begins at a single origin. That setup creates two replication forks that move in opposite directions from the opening...

proofreading

Proofreading in DNA replication helps keep DNA copying accurate. It is a built-in check that reduces mistakes as new DNA is made. DNA polymerase is the enzyme that carries out this checking during replication. When it adds a wrong nucleotide, it can remove the error and replace it with the correct one. This correction step helps maintain the correct DNA sequence. Proofreading is an important part of DNA replication because it supports genetic accuracy. By catching mistakes as they happen, the...

dna helicases

Helicase Motor Proteins in DNA Unwinding

Helicase motor proteins help unwind DNA during key cell processes. They use energy from ATP hydrolysis, which is the breakdown of ATP to release usable energy. This movement lets helicases separate DNA strands when the cell needs access to the genetic code. These enzymes are involved in DNA replication, repair, recombination, and transcription. Helicases are found in all living organisms, but their structure, function, and movement can differ. In prokaryotes, the DnaB helicase moves along the...

the replisome

DNA Replication Teams in the Replisome

DNA replication depends on the replisome, a coordinated complex of proteins that copies DNA with high accuracy. This DNA replication machinery brings many parts together so the leading and lagging strands can be made at the same time. A key idea in this process is the trombone model, proposed by Bruce Alberts in 1980. In this model, a loop forms on the lagging strand when a primer is made. The loop grows as an Okazaki fragment is synthesized, then falls apart when that fragment is finished.

lagging strand synthesis

Okazaki Fragments in DNA Replication

DNA replication uses a leading strand and a lagging strand, and they are made at different rates. The leading strand starts first. The lagging strand starts later, moves more slowly, and is made in pieces. The two strands also differ in direction and starting steps. Leading strand synthesis follows the opening replication fork. Lagging strand synthesis goes in the opposite direction. The leading strand needs one RNA primer, while the lagging strand needs multiple RNA primers. After priming,...

mismatch repair

Fixing DNA Copying Errors

Mismatch repair helps fix DNA copying errors after replication. When DNA is copied, small mistakes can slip through. This repair system finds those mismatches and corrects them before they become permanent changes. The process depends on a group of mutator proteins. These proteins work together to detect the error, remove the incorrect section, and help replace it with the proper DNA sequence. In this way, mismatch repair supports the accuracy of genetic information. If the repair system does...

dna topoisomerases

How Topoisomerases Untwist DNA

DNA topoisomerases are enzymes that untwist DNA when it becomes overcoiled during cell processes such as DNA replication and transcription. They help regulate positive and negative DNA supercoiling without changing the nucleotide sequence. Clockwise overwinding creates positively supercoiled DNA, while counterclockwise underwinding produces negatively supercoiled DNA. Topoisomerases are grouped into two main types. Type I topoisomerases act on one strand of double-stranded DNA, and they...

telomeres and telomerase

How Telomerase Protects Chromosome Ends

Telomerase helps protect chromosome ends during eukaryotic DNA replication. When the final primer is removed, a single-stranded DNA fragment can remain at the end of the chromosome. That piece cannot be copied in the same way as the rest of the strand because there is no 3' end for new DNA to attach to. If this end piece is not replaced, chromosomal DNA is gradually lost each time a cell divides. The exposed single-stranded DNA can also trigger a DNA damage response from enzymes that detect it.

non-nuclear inheritance

Cytoplasmic Inheritance in Cells

Cytoplasmic inheritance explains how some traits are passed through cell structures outside the nucleus. These traits come from organelles such as mitochondria and chloroplasts. They do not follow the usual nuclear pattern of inheritance. Mitochondria and chloroplasts each contain their own DNA. Because of this, they can carry genetic information that affects cell function and certain visible traits. In high school biology, this topic helps explain why not all inherited traits come from...

animal mitochondrial genetics

Animal Mitochondrial DNA and Inheritance

Animal mitochondrial DNA has its own genome, separate from nuclear DNA. It is a double-stranded, closed circular molecule with about 20,000 base pairs. One strand is the heavy, or H-strand, because it is rich in guanine. The other is the light, or L-strand, because it is rich in cytosine. Compared with nuclear DNA, mitochondrial DNA has very little non-coding sequence. It also has no introns. Its genes are packed closely together, and some even overlap. The D-loop is the main non-coding...

comparing mitochondrial, chloroplast, and prokaryotic genomes

comparing-mitochondrial-chloroplast-and-prokaryotic-genomes

How Organelle Genomes Resemble Bacteria

Mitochondrial and chloroplast genomes still keep several traits from their bacterial ancestors. They also show changes that happened after these organelles evolved inside eukaryotic cells. Eukaryotic cells are cells with a nucleus, while prokaryotes are cells without one. Like prokaryotic genomes, mitochondrial and chloroplast genomes do not bind with histone-like proteins. They also do not show the complex chromosome-like packaging seen in eukaryotes. Mitochondria and chloroplasts divide by...

export of mitochondrial and chloroplast genes

export-of-mitochondrial-and-chloroplast-genes

How Organelles Pass Genes to the Nucleus

Eukaryotic cells can carry three genetic systems: nuclear, mitochondrial, and chloroplast. Over time, many genes from mitochondria and chloroplasts have moved into the nucleus. In some plant species, this gene transfer is still happening. This movement has affected large parts of the genome in some organisms. About 18% of the Arabidopsis thaliana nuclear genome is thought to come from the chloroplast's cyanobacterial ancestor. In yeast, around 75% of the genome is derived from the...

single-strand dna binding proteins

How SSB Proteins Protect DNA Strands

Single-strand DNA-binding proteins, or SSB proteins, help protect DNA during replication. When double-stranded DNA is unwound, the two strands separate. SSB proteins bind to the exposed single strands and keep them stable. These proteins bind in a sequence-independent manner. That means the order of the nitrogenous bases does not have to follow a specific pattern for SSB proteins to attach. Their binding straightens single-stranded DNA and makes it more rigid. This support may help DNA...

key terms and concepts

Learn how proofreading works during DNA replication and how DNA polymerase helps correct mistakes to keep the DNA sequence accurate.

Proofreading in DNA Replication | JoVE K12

molecular biology

JoVE Core Biologie Moléculaire illustre les processus moléculaires au sein de la cellule grâce à des leçons vidéos claires et concises. Les applications de ces concepts sont ensuite démontrées grâce à des expériences concrètes en laboratoire.

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Proofreading in DNA Replication

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