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Overview of DNA Replication

DNA replication is a vital process for genetic material duplication before cell division, involving initiation, elongation, and termination phases. It starts at specific origins with the help of proteins like DnaA in prokaryotes and the ORC in eukaryotes. The pre-replication complex formation is crucial for the accurate and regulated commencement of replication. DNA polymerases, helicases, and other enzymes work together at the replication fork to synthesize new DNA strands, while telomerase resolves the end-replication problem in eukaryotes.

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1

Role of AT-rich regions in DNA replication initiation

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AT-rich regions have fewer hydrogen bonds, making DNA strands easier to separate for replication initiation.

2

Function of initiator proteins in prokaryotes vs. eukaryotes

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In prokaryotes, DnaA binds to the origin; in eukaryotes, ORC binds to start DNA replication.

3

Purpose of the pre-replication complex

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Ensures DNA replication starts accurately and occurs only once per cell cycle, preventing DNA over-replication.

4

In eukaryotic cells, the ______ recruits proteins like Cdc6 and Cdt1 to load the ______ helicase onto DNA.

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origin recognition complex (ORC) minichromosome maintenance (Mcm)

5

The transition from ______ to ______ phase and the start of DNA synthesis are triggered by ______ and ______ kinase.

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G1 S cyclin-dependent kinases (Cdks) Dbf4-dependent kinase (DDK)

6

Function of Mcm helicase in DNA replication

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Mcm helicase unwinds DNA double helix at replication origins, enabling strand separation for template reading.

7

Role of primase in DNA synthesis

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Primase synthesizes short RNA primers to provide starting points for DNA polymerase on both leading and lagging strands.

8

Synthesis difference between leading and lagging strands

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Leading strand synthesized continuously towards replication fork; lagging strand synthesized in Okazaki fragments, later joined by DNA ligase.

9

In the process of DNA replication, nucleotides are added in a specific direction, which is the ______ to ______ direction.

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5′ 3′

10

During DNA replication in prokaryotes, ______ is the main enzyme for replication, while ______ removes RNA primers.

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DNA polymerase III DNA polymerase I

11

Function of SSBs at replication fork

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SSBs stabilize unwound DNA, preventing re-annealing.

12

Synthesis direction of leading vs lagging strands

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Leading strand synthesized continuously, lagging strand discontinuously.

13

Purpose of Okazaki fragments

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Enable synthesis of lagging strand in looped configuration.

14

Proteins such as DNA ______, ______, sliding clamps, and others are part of the replisome ensuring accurate DNA replication.

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helicases polymerases

15

During DNA replication, replisome components are ______ regulated, allowing them to assemble and disassemble as necessary.

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dynamically

16

In ______ cells, DNA replication occurs at multiple origins and is coordinated by replisomes to complete before ______ division.

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eukaryotic cell

17

End-replication problem in eukaryotes

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Eukaryotic linear chromosomes' terminal regions can't be fully replicated by DNA polymerases, leading to shorter telomeres after each division.

18

Function of telomerase

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Telomerase extends telomeres in eukaryotes to preserve genetic information and counteract the end-replication problem.

19

Termination of replication in prokaryotes

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Prokaryotic circular chromosomes use specific sequences and proteins to ensure replication stops at the correct site, avoiding over-replication.

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Overview of DNA Replication

DNA replication is a critical biological process that ensures the accurate duplication of the genetic material prior to cell division. This process is highly conserved and involves a series of coordinated steps: initiation, elongation, and termination. During initiation, specific proteins recognize and bind to origins of replication, which are particular sequences in the DNA where replication begins. In prokaryotes, such as E. coli, the initiator protein DnaA binds to the origin, whereas in eukaryotes, the origin recognition complex (ORC) performs a similar function. These proteins facilitate the unwinding of the DNA double helix, which is rich in adenine-thymine (AT) pairs at these sites due to their lower number of hydrogen bonds, making them easier to separate. The formation of the pre-replication complex at these origins ensures that DNA replication commences accurately and is strictly regulated to occur only once during each cell cycle, preventing the duplication of DNA segments.
Three-dimensional model of double helix DNA with base pairs colored blue and red for adenine and thymine, green and yellow for cytosine and guanine, on a blurred laboratory background.

Formation and Regulation of the Pre-replication Complex

The assembly of the pre-replication complex is a critical event that takes place during the late mitosis and early G1 phase of the cell cycle. This complex is composed of a multitude of proteins that prepare the DNA for replication. In eukaryotic cells, the origin recognition complex (ORC) recruits additional proteins such as Cdc6 and Cdt1, which are instrumental in loading the minichromosome maintenance (Mcm) helicase onto the DNA. This helicase is essential for unwinding the DNA at the replication forks. The activation of the pre-replication complex is tightly regulated by cyclin-dependent kinases (Cdks) and Dbf4-dependent kinase (DDK), which includes Cdc7, crucial for the transition from G1 to S phase and the commencement of DNA synthesis. These regulatory mechanisms ensure that DNA replication is precisely coordinated with the cell cycle and adapts to the cellular environment.

The Preinitiation Complex and DNA Synthesis

The transition into the S phase of the cell cycle triggers the formation of the preinitiation complex, which activates the Mcm helicase, leading to the unwinding of the DNA double helix. This complex also recruits DNA polymerases to the replication origins. DNA synthesis begins with the synthesis of short RNA primers by primase, which provides a starting point for DNA polymerases. The leading strand is synthesized continuously in the direction of the replication fork, while the lagging strand is synthesized in short, discontinuous segments known as Okazaki fragments. These RNA primers are subsequently replaced with DNA, and the Okazaki fragments are ligated together by DNA ligase to produce a continuous strand, completing the replication process.

Mechanisms of DNA Strand Elongation

DNA polymerases catalyze the elongation of new DNA strands by adding nucleotides in a 5′ to 3′ direction. The leading strand is synthesized continuously, while the lagging strand is synthesized in segments, which are later connected to form a complete strand. In prokaryotes, DNA polymerase III is the primary enzyme for replication, with DNA polymerase I playing a role in removing RNA primers and replacing them with DNA. In eukaryotes, DNA polymerase alpha (Pol α) initiates DNA synthesis, while DNA polymerase epsilon (Pol ε) and DNA polymerase delta (Pol δ) are responsible for the elongation of the leading and lagging strands, respectively. The primase used in eukaryotes and archaea is structurally distinct from that in bacteria, highlighting the evolutionary diversity of replication mechanisms.

The Dynamics of the Replication Fork

The replication fork is a dynamic structure where the DNA double helix is unwound to allow the copying of template strands. DNA helicases are responsible for unwinding the DNA, and single-strand binding proteins (SSBs) stabilize the unwound DNA to prevent it from re-annealing. Topoisomerases alleviate the torsional stress that accumulates ahead of the replication fork, preventing supercoiling and potential DNA damage. The replication fork is the site of synthesis for both the leading and lagging strands, with the latter requiring a looped configuration to enable the synthesis of Okazaki fragments.

DNA Replication Proteins and the Replisome

The replisome is a complex assembly of proteins that collaborate at the replication fork to ensure the fidelity and efficiency of DNA replication. This includes DNA helicases, polymerases, sliding clamps, clamp loaders, and single-strand binding proteins, among others. The replisome components are dynamically regulated, assembling and disassembling as needed throughout the replication process. In eukaryotic cells, replication occurs simultaneously at multiple origins along the chromosomes, and the replisomes coordinate these processes to ensure that replication is completed accurately and in a timely manner before cell division.

Termination of DNA Replication

DNA replication concludes when replication forks converge, and the newly synthesized DNA strands are fully elongated. In eukaryotes, this can occur at various termination sites due to the presence of multiple origins of replication. The linear structure of eukaryotic chromosomes presents the end-replication problem, where the terminal regions, known as telomeres, cannot be completely replicated by conventional DNA polymerases. The enzyme telomerase extends these regions to preserve genetic information. In prokaryotes, which typically have circular chromosomes, termination is facilitated by specific sequences and termination proteins that ensure replication ceases at the appropriate site. Proper regulation of termination is essential for maintaining genomic stability and ensuring accurate cell division.