Name: dna replication teams in the replisome
Uploaded: 2020-11-23T11:47:35.000Z
Duration: 3 min 1 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

The Replisome

进行DNA复制 高度协调 多蛋白组装称为 DNA复制机制，或复制品，提高DNA效率 复制。核心组成 机械的 是解旋酶，单链 DNA结合蛋白 DNA primase，滑动夹，钳式装载器和多个DNA 聚合酶，都是 彼此关联 复制叉附近。复制性DNA聚合酶 有生产力 大约有10个核苷酸 是核苷酸的数量 可以在解离前 加到女链上 从模板链。这太低效了 复制整个基因组 在合理的时间内 框架，这个问题 在帮助下解决了 滑动钳蛋白。当ATP与 钳式装载机蛋白 他们将绑定到和 打开滑动夹，使其呈环状 结构可以 包围 引物-模板DNA复合物。一旦绑定，钳 装载机水解ATP 到ADP，造成钳位 装载机解除关联 和钳 紧紧围绕着DNA。然后DNA聚合酶结合 夹蛋白。一起滑 沿着模板DNA，束缚DNA 聚合酶链 并提高其生产率 最多1, 000个核苷酸。生产力的提高 允许DNA聚合酶 进行连续的DNA 复制领先 股。但是，在滞后 链模板，另一个DNA聚合酶执行 不连续的DNA复制 以允许的方式 DNA聚合酶 要合成的分子 领先和落后 同时绞合。这个过程是 有时描述 作为长号模型。滞后的钢绞线 及其模板 当形成绞合线时 DNA聚合酶引发 冈崎片段合成 从RNA引物。DNA环在增长 从两个方向 当解旋酶解开DNA和 滞后链被合成。当DNA聚合酶 遇到下一个RNA引物，它脱离 模板链。同时，primase添加了另一个 落后链的引物。以及不断增长的DNA 循环被释放。夹钳和夹钳 装载蛋白 允许DNA聚合酶 快速重新关联 带有引物的DNA模板。形成及后续 DNA环的崩溃 用合成重复 每个新的冈崎片段。

<ol>
	<li>O'Donnell、Michael、Lance Langston 和 Bruce Stillman。"细菌、古细菌和真核生物中 DNA 复制的原理和概念。"冷泉港生物学观点 5，第 7 期（2013 年）:a010108。</li>
	<li>阿尔伯茨，布鲁斯。"DNA 复制和重组。"自然 421，第 6921 期（2003）:431。</li>
	<li>Kelch、Brian A.、Debora L. Makino、Mike O'Donnell 和 John Kuriyan。"夹式装载机 ATP 酶和 DNA 复制机制的进化。"BMC 生物学 10，第 1 期（2012）:34。</li>
	<li>Hedglin、Mark、Ravindra Kumar 和 Stephen J. Benkovic。"复制夹具和夹具装载机。"冷泉港生物学观点 5，第 4 期（2013 年）:a010165。</li>
	<li>加德纳、安德鲁 F. 和 Zvi 凯尔曼。"作为治疗靶点的 DNA 复制机制。"分子生物科学前沿 6 （2019）:35。</li>
</ol>

复制体

DNA 复制是由大量蛋白复合体进行的，这些蛋白以协调的方式作用，以实现高保真 DNA 复制。 这个复合体一起被称为 DNA 复制机器或复制体。

前导链和滞后链的合成是一个高度协调的过程。 为了解释这一点，Bruce Alberts 于 1980 年提出了“长号模型”。当在亲本滞后链上合成引物时，DNA 环的形成就开始了。 该循环随着冈崎片段的合成而增长，并最终在片段合成结束时溶解。 然后 DNA 聚合酶附着到另一个引物上，并重复从环形成到塌陷的整个循环。 复制体允许所有活动作为复制机器复合体进行协调。

复制体组件

解旋酶解旋并分离双链 DNA。 这些酶在原核生物中以单六聚体环存在，在真核生物中以双六聚体环存在。 真核解旋酶依赖于额外的蛋白 Cdc45 和 GINS 来发挥作用。 单链 DNA 结合 (SSB) 蛋白可防止 DNA 链重新退火。 在原核生物中，SSB 由单个亚基组成，而在真核生物中，它们是一种异三聚体蛋白，称为复制蛋白 A (RPA)。

Primase 在 DNA 合成起始处添加 RNA 引物。 在原核生物中，引物酶以称为 DnaG 的单亚基酶形式存在，它合成约 12 个核苷酸的 RNA 引物。 在真核生物中，一种多亚基酶 DNA 聚合酶-α 引物酶可合成约 25 个核苷酸的 RNA-DNA 杂合引物。 除了聚合酶-α 之外，复制聚合酶也会延伸新合成的 DNA。 原核生物中存在单一类型的复制聚合酶 DNA 聚合酶 III，而真核生物中存在两种不同类型的复制聚合酶 Pol ε 和 Pol δ，分别用于前导链和滞后链的合成。

滑动钳蛋白使聚合酶附着在 DNA 模板上。 β-clamp 是一种同二聚体蛋白，在原核生物中充当钳子的作用。 在真核生物中，同样的任务是由增殖细胞核抗原（PCNA）（一种同源三聚体蛋白）完成的。 滑动钳蛋白的中心孔带正电，这使其能够与带负电的 DNA 磷酸盐主链发生静电相互作用。 通过钳子装载器将钳子连接到 DNA 上。 这些五聚体蛋白属于 AAA+ 类 ATP 酶。 真核夹钳装载机被称为复制因子C，而原核大肠杆菌夹钳装载机被称为γ复合体； 然而，与复制体的许多其他成分不同，钳蛋白和钳加载器被认为是进化同源物。

DNA复制由一大型蛋白质复合物执行，这些蛋白质以协调的方式共同作用，实现高保真的DNA复制。该复合物协同工作，共同完成……

Education

教育

JoVE Core

Molecular Biology

分子生物学

Unit 1

第一单元

DNA Replication

DNA 复制

KEY TERMS AND CONCEPTS

关键术语与概念

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. DNA polymerase then moves to a new primer, and the cycle repeats.

Helicases start the process by unwinding and separating double-stranded DNA. In prokaryotes, helicase is a single hexamer ring, while in eukaryotes it forms a double hexamer ring. Eukaryotic helicases also need the extra proteins Cdc45 and GINS to work. Single-stranded DNA binding proteins keep the separated strands from reannealing. In prokaryotes, these are SSB proteins with one subunit. In eukaryotes, the equivalent protein is replication protein A, or RPA, which is a heterotrimer.

Primase adds the RNA primer that gives DNA synthesis a starting point. In prokaryotes, primase is the single-subunit enzyme DnaG, and it makes an RNA primer of about 12 nucleotides. In eukaryotes, DNA polymerase-alpha primase makes an RNA-DNA hybrid primer of about 25 nucleotides. After that, the replicative polymerase extends the new DNA strand. Prokaryotes use DNA polymerase III, while eukaryotes use Pol epsilon for the leading strand and Pol delta for the lagging strand.

Sliding clamp proteins hold the polymerases on the DNA template so copying can continue efficiently. The prokaryotic beta-clamp is a homodimer, and the eukaryotic clamp is proliferating cell nuclear antigen, or PCNA, which is a homotrimer. The clamp has a positively charged central pore that interacts with the negatively charged DNA backbone. A clamp loader places the clamp onto DNA. These pentameric AAA+ ATPases include the eukaryotic replication factor C and the E. coli gamma complex. The clamp proteins and clamp loaders are thought to be evolutionary homologs, unlike many other replisome components.

DNA Replication Teams in the Replisome

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

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 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

Explore how the replisome coordinates DNA replication, from helicase and primase to polymerases, sliding clamps, and the trombone model.

DNA Replication Teams in the Replisome | JoVE K12

molecular biology

JoVE Core 分子生物学通过清晰简洁的动画视频课程展示了细胞的分子过程。此外，在科学家在行动的视频中，这些概念的应用是通过现代实验室中的真实实验演示的。

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1. Engage in evidence-based argument to relate a cell's function to the structure, function, and diversity of its components. Examples: Muscle cells have a large number of mitochondria, which create energy for contraction in the form of ATP. Some plant cells have a large vacuole which is used for storage and transport of excess nutrients in changing environments.

9-12.B.1.1

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Types of RNA

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Transcription

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Bacterial RNA Polymerase

Eukaryotic RNA Polymerases

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Transcription Elongation Factors

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Pre-mRNA Processing: RNA Splicing

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Alternative RNA Splicing

Nuclear Export of mRNA

Transfer RNA Synthesis

Ribosomal RNA Synthesis

The Nucleolus

Additional Subnuclear Structures

Translation

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tRNA Activation

Initiation of Translation

Termination of Translation

Improving Translational Accuracy

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Nonsense-mediated mRNA Decay

The Eukaryotic Promoter Region

Molecular Chaperones and Protein Folding

Co-activators and Co-repressors

The Proteasome

Master Transcription Regulators

Regulated Protein Degradation

Regulated mRNA Transport

Proteins: From Genes to Degradation

mRNA Stability and Gene Expression

Cell Specific Gene Expression

MicroRNAs

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a. Use a model to describe the structure and sequence of DNA, including nucleotide structure, base pairing, and the structure of the helix.

9-12.B.1.2.a

DNA Replication Teams in the Replisome

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