4.3 Replication in prokaryotes and eukaryotes
4 min read•august 16, 2024
DNA replication is a crucial process for life, copying genetic material before cell division. Prokaryotes and eukaryotes share basic principles but differ in complexity and regulation.
Prokaryotes have simpler, faster replication with one origin on a circular chromosome. Eukaryotes have multiple origins on linear chromosomes, deal with chromatin, and have stricter cell cycle control.
DNA Replication in Prokaryotes vs Eukaryotes
Fundamental Similarities and Differences
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- applies to both prokaryotic and generates new double helix with one original strand and one newly synthesized strand
- occurs in cytoplasm while eukaryotic DNA replication takes place in nucleus
- Prokaryotes typically have single circular chromosome with one origin of replication
- Eukaryotes possess multiple linear chromosomes with multiple origins of replication
- DNA replication speed in prokaryotes reaches approximately 1000 nucleotides/second
- Eukaryotic DNA replication proceeds at slower rate of approximately 50 nucleotides/second
- Eukaryotic DNA replication involves additional complexities
- Chromatin structure management
- Presence of at chromosome ends
- Prokaryotic DNA replication couples with cell division
- Eukaryotic DNA replication restricts to of cell cycle
Structural and Functional Complexities
- Eukaryotic chromosomes contain histone proteins forming nucleosomes
- Nucleosomes must be disassembled and reassembled during replication
- Eukaryotic telomeres require specialized mechanisms for replication
- Telomerase enzyme adds repetitive sequences to chromosome ends
- Prokaryotic circular chromosome forms supercoiled structure
- relieve DNA tension during replication
- Eukaryotic replication involves larger number of proteins and regulatory factors
- act as replicative helicases
- serves as sliding clamp for DNA polymerases
Origin of Replication in DNA Synthesis
Structure and Recognition of Origins
- consists of specific DNA sequence initiating replication
- Prokaryotic origin recognition
- binds to specific sequences called
- DnaA boxes typically contain 9-base pair consensus sequence (TTATCCACA)
- Eukaryotic origin recognition
- binds to replication origins
- ORC composed of six subunits (ORC1-6)
- Eukaryotic origins often associated with AT-rich sequences
Initiation Complex Formation
- Binding of initiator proteins to origin leads to local unwinding of DNA double helix
- Replication bubble forms at unwound region
- Multiple proteins recruited to origin form
- Prokaryotes: loaded onto DNA by DnaC protein
- Eukaryotes: loaded by Cdc6 and proteins
- Helicases further unwind DNA preparing template for replication
- Number and distribution of origins affect overall replication timing and efficiency
- Prokaryotes typically have single origin (oriC in E. coli)
- Eukaryotes contain thousands of origins spaced along chromosomes
Significance of the Replication Fork
Structure and Components of the Replication Fork
- forms Y-shaped region where DNA strands separate for new strand synthesis
- Key components of replication fork
- DNA helicase unwinds parental DNA strands
- synthesizes short RNA primers (5-10 nucleotides long)
- initiates synthesis from 3' hydroxyl group of primers
- Single-stranded binding proteins (SSB) stabilize unwound single-stranded DNA
- synthesized continuously in 5' to 3' direction
- synthesized discontinuously as
- Okazaki fragments typically 100-200 nucleotides long in prokaryotes
- Eukaryotic Okazaki fragments shorter (approximately 100-150 nucleotides)
Enzymatic Activities at the Replication Fork
- DNA polymerase III (prokaryotes) or DNA polymerase δ and ε (eukaryotes) catalyze nucleotide addition
- joins Okazaki fragments on lagging strand
- removes RNA primers
- (prokaryotes) or DNA polymerase δ (eukaryotes) fills gaps left by primer removal
- Topoisomerases relieve DNA supercoiling ahead of replication fork
- Prokaryotes use DNA gyrase (type II topoisomerase)
- Eukaryotes employ topoisomerase I and II
Regulation of DNA Replication: Prokaryotes vs Eukaryotes
Prokaryotic Replication Regulation
- Prokaryotic replication primarily regulated at initiation stage
- DnaA protein availability and activity control replication initiation
- DnaA-ATP active form binds origin, while DnaA-ADP inactive
- Regulatory mechanisms in prokaryotes
- Sequestration of newly replicated origins by SeqA protein (E. coli)
- Titration of DnaA protein by multiple DnaA binding sites across genome
- Regulation of DnaA gene expression
- Prokaryotes adjust replication rate in response to nutrient availability
- Nutrient-rich conditions increase initiation frequency
Eukaryotic Replication Regulation
- Eukaryotic replication tightly controlled throughout cell cycle
- Two-step process for origin activation
- Licensing of origins in G1 phase
- Activation of licensed origins in S phase
- play crucial role in regulating replication
- CDK activity low in G1 allows origin licensing
- CDK activity high in S phase promotes origin firing and prevents re-licensing
- Multiple mechanisms prevent re-replication within single cell cycle
- Degradation of licensing factors (Cdt1)
- Nuclear export of MCM proteins
- Inactivation of origins after firing
- Replication timing regulation in eukaryotes
- Concept of early and late-replicating domains
- Influenced by chromatin structure and nuclear organization
- Transcriptionally active regions often replicate early
Key Terms to Review (29)
Cdt1: Cdt1 (Chromatin licensing and DNA replication factor 1) is a protein essential for the initiation of DNA replication in eukaryotic cells. It functions by ensuring that DNA replication occurs only once per cell cycle by helping to load the MCM (Minichromosome Maintenance) complex onto DNA during the G1 phase. This regulation is crucial for maintaining genomic stability and preventing re-replication within a single cell cycle.
Cyclin-dependent kinases (cdks): Cyclin-dependent kinases (cdks) are a family of protein kinases that play a crucial role in regulating the cell cycle by phosphorylating specific target proteins. They are activated by binding to cyclins, which are regulatory proteins that fluctuate in concentration throughout the cell cycle, ensuring that cdks function at the right time. This precise regulation allows for proper DNA replication, mitosis, and ultimately, cell division.
Dna ligase: DNA ligase is an essential enzyme that facilitates the joining of DNA strands together by catalyzing the formation of phosphodiester bonds. This process is crucial during DNA replication and repair, where it seals nicks in the sugar-phosphate backbone of DNA, ensuring the integrity and continuity of the genetic material. Without DNA ligase, proper replication and repair processes would be severely hindered, leading to potential mutations and genomic instability.
Dna polymerase: DNA polymerase is an essential enzyme that synthesizes new DNA strands by adding nucleotides to a growing DNA chain during replication and repair processes. This enzyme plays a critical role in ensuring the accuracy and efficiency of DNA replication, facilitating various applications such as amplification in laboratory techniques, addressing DNA damage, and functioning in both prokaryotic and eukaryotic organisms.
Dna polymerase i: DNA polymerase I is an enzyme crucial for DNA replication, primarily known for its role in prokaryotic cells. It is responsible for synthesizing new DNA strands by adding nucleotides to the growing chain and has 5' to 3' polymerase activity, along with 3' to 5' exonuclease activity for proofreading. Its function is vital in the processing of Okazaki fragments during lagging strand synthesis, helping to ensure fidelity in DNA replication.
Dna primase: DNA primase is an essential enzyme that synthesizes short RNA primers during DNA replication, providing a starting point for DNA polymerase to extend the new DNA strand. By creating these primers, primase plays a critical role in both prokaryotic and eukaryotic DNA replication processes, ensuring that the replication machinery functions efficiently and accurately.
DnaA boxes: dnaA boxes are specific DNA sequences found within the origin of replication in prokaryotic organisms, primarily serving as binding sites for the DnaA protein, which is essential for initiating DNA replication. These sequences play a critical role in the replication process by helping to unwind the DNA helix and recruit other proteins necessary for the formation of the replication complex. The proper functioning of dnaA boxes is crucial for the timely and accurate replication of prokaryotic genomes.
Dnaa protein: Dnaa protein is a vital initiator protein in prokaryotic DNA replication, responsible for recognizing and binding to the origin of replication, known as the DnaA box. Once bound, it facilitates the unwinding of the DNA double helix, allowing other proteins involved in replication to access the single-stranded DNA. This process is essential for initiating the replication fork and ensuring the proper duplication of genetic material in bacterial cells.
Dnab helicase: DnaB helicase is an essential enzyme in prokaryotic DNA replication that unwinds the DNA double helix at the replication fork, allowing the two strands to be separated and accessible for copying. This enzyme operates in conjunction with other proteins and factors to ensure efficient and accurate DNA replication. Its role is crucial for both the initiation and elongation phases of replication in bacteria.
Eukaryotic dna replication: Eukaryotic DNA replication is the process by which a eukaryotic cell duplicates its genetic material in preparation for cell division. This complex process involves multiple origins of replication, the formation of a replication fork, and several key enzymes, ensuring the accurate copying of the entire genome, which is organized into linear chromosomes.
Lagging strand: The lagging strand is one of the two strands of DNA being synthesized during DNA replication, characterized by its discontinuous synthesis in short segments known as Okazaki fragments. This strand is formed in the opposite direction of the replication fork movement and requires multiple RNA primers to initiate each fragment. Understanding its formation is crucial for grasping the overall process of DNA replication and the structural properties of DNA.
Leading strand: The leading strand is the continuously synthesized DNA strand during DNA replication that runs in the 5' to 3' direction, mirroring the unwound template strand. It is synthesized by DNA polymerase as the replication fork progresses, allowing for a smooth and efficient replication process. Understanding the leading strand is essential to grasping the intricacies of DNA replication mechanics and its differences between prokaryotic and eukaryotic organisms.
Mcm2-7 helicase complex: The mcm2-7 helicase complex is a crucial protein assembly in eukaryotic cells that functions to unwind DNA strands during the replication process. This hexameric complex is composed of six subunits, which form a ring structure that encircles the DNA, allowing it to move along the double helix and separate the strands ahead of the replication fork. Its activity is essential for the proper initiation and elongation of DNA replication, highlighting its role in ensuring genomic stability and accurate DNA synthesis.
Minichromosome maintenance (mcm) proteins: Minichromosome maintenance (mcm) proteins are essential helicase enzymes that play a critical role in the initiation of DNA replication. They help to unwind and separate the DNA strands, allowing for the replication machinery to access the DNA template. These proteins are found in both prokaryotic and eukaryotic organisms, but their roles and regulation can vary significantly between these two domains of life.
Okazaki Fragments: Okazaki fragments are short sequences of DNA that are synthesized on the lagging strand during DNA replication. These fragments are essential because they allow for the discontinuous synthesis of DNA, which occurs due to the antiparallel nature of DNA strands and the directionality of DNA polymerase. Understanding Okazaki fragments helps clarify the overall process of DNA replication, including how it differs in prokaryotes and eukaryotes, as well as the enzymes involved in this critical cellular function.
Origin of replication (ori): The origin of replication (ori) is a specific sequence of nucleotides where DNA replication begins. It plays a critical role in both prokaryotic and eukaryotic cells by determining how the DNA is copied during cell division, ensuring that each daughter cell receives an exact copy of the genetic material.
Origin Recognition Complex (ORC): The Origin Recognition Complex (ORC) is a multi-subunit protein complex that plays a critical role in the initiation of DNA replication in eukaryotic cells. It binds to specific sites on the DNA, known as origins of replication, and serves as a platform for recruiting other proteins necessary for the replication process. The function of ORC is essential for ensuring that DNA replication occurs accurately and at the right time during the cell cycle.
Pre-replication complex (pre-rc): The pre-replication complex (pre-rc) is a protein assembly that forms at the origin of replication during the preparation phase for DNA replication. This complex is essential for ensuring that DNA replication occurs accurately and efficiently, as it includes several key proteins that facilitate the unwinding of the DNA double helix and the recruitment of DNA polymerases necessary for synthesis. The pre-rc plays a critical role in both prokaryotic and eukaryotic organisms by regulating the timing and initiation of DNA replication.
Prokaryotic dna replication: Prokaryotic DNA replication is the process by which a single, circular DNA molecule in prokaryotic cells is duplicated to ensure that each daughter cell receives an identical copy during cell division. This process is characterized by its speed and simplicity compared to eukaryotic DNA replication, involving fewer proteins and a more streamlined mechanism due to the lack of membrane-bound organelles.
Proliferating Cell Nuclear Antigen (PCNA): Proliferating Cell Nuclear Antigen (PCNA) is a protein that acts as a processivity factor for DNA polymerase during DNA replication, essentially helping to clamp the enzyme onto the DNA strand. This allows for more efficient and accurate DNA synthesis in both prokaryotic and eukaryotic cells, linking it directly to the fundamental processes of cell division and repair mechanisms.
Replication checkpoint: A replication checkpoint is a regulatory mechanism that ensures the integrity and accuracy of DNA replication before the cell proceeds to division. This checkpoint assesses whether DNA is fully and correctly replicated, allowing for any necessary repairs to be made, which prevents the propagation of mutations and maintains genomic stability.
Replication Fork: A replication fork is a Y-shaped structure that forms during DNA replication, where the double-stranded DNA molecule unwinds and separates into two single strands. This structure is crucial for the process of DNA duplication, allowing enzymes to access the single-stranded DNA to synthesize new complementary strands. The replication fork facilitates both leading and lagging strand synthesis, and is a key feature in understanding how DNA is replicated in both prokaryotes and eukaryotes.
Replication licensing: Replication licensing is a crucial regulatory mechanism that ensures DNA is only replicated once during each cell cycle. This process involves the assembly of specific proteins at the origins of replication, allowing for the controlled initiation of DNA synthesis in both prokaryotic and eukaryotic cells. Proper licensing is essential to prevent re-replication, which can lead to genomic instability and other cellular problems.
RNase H: RNase H is an enzyme that specifically degrades the RNA strand of RNA-DNA hybrids. It plays a crucial role in various biological processes, including DNA replication and repair, by removing RNA primers during DNA synthesis, thereby facilitating the transition from RNA to DNA. This function is essential for maintaining genomic integrity in both prokaryotes and eukaryotes, highlighting its importance in the overall process of DNA replication.
S Phase: The S phase, or synthesis phase, is a crucial part of the cell cycle during which DNA replication occurs. During this phase, the genetic material of the cell is duplicated to ensure that each daughter cell receives an identical copy of the genome after cell division. This phase is essential for both prokaryotic and eukaryotic organisms, as it sets the foundation for accurate cell division and genetic continuity.
Semi-conservative model: The semi-conservative model describes the mechanism of DNA replication where each of the two strands of the original DNA molecule serves as a template for the formation of new complementary strands. This means that after replication, each new double helix consists of one old strand and one newly synthesized strand, ensuring genetic continuity and stability across generations.
Single-stranded binding proteins (ssbs): Single-stranded binding proteins (ssbs) are proteins that bind to single-stranded DNA during the processes of DNA replication and repair, stabilizing the unwound DNA strands. Their primary role is to prevent the re-annealing of the separated strands and protect them from degradation by nucleases, ensuring that the replication process proceeds smoothly and accurately. ssbs are crucial in both prokaryotic and eukaryotic organisms, as they help maintain the integrity of the genetic material during replication.
Telomeres: Telomeres are repetitive nucleotide sequences located at the ends of linear chromosomes, which protect them from deterioration or fusion with neighboring chromosomes. They play a crucial role in maintaining genomic stability and are involved in cellular aging, as they shorten with each cell division, ultimately leading to cellular senescence when they become critically short.
Topoisomerases: Topoisomerases are essential enzymes that regulate the supercoiling of DNA during processes like replication and transcription. They introduce temporary breaks in the DNA strands, allowing the DNA to unwind or rewind, thereby alleviating the tension created by the winding and unwinding of the double helix. By managing these topological changes, they play a crucial role in ensuring that DNA can be accurately replicated and transcribed in both prokaryotic and eukaryotic cells.