Steps of DNA Replication

01
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The Process of DNA Replication

DNA Replication
DNA replication faithfully duplicates the entire genome of the cell. During DNA replication, a number of different enzymes work together to pull apart the two strands so each strand can be used as a template to synthesize new complementary strands. OpenStax, Anatomy & Physiology/Creative Commons Attribution 3.0

Why Replicate DNA?

DNA is the genetic material that defines every cell. Before a cell duplicates and is divided into new daughter cells through either mitosis or meiosis, biomolecules and organelles must be copied to be distributed among the cells. DNA, found within the nucleus, must be replicated in order to ensure that each new cell receives the correct number of chromosomes. The process of DNA duplication is called DNA replication. Replication follows several steps that involve multiple proteins called replication enzymes and RNA. In eukaryotic cells, such as animal cells and plant cells, DNA replication occurs in the S phase of interphase during the cell cycle. The process of DNA replication is vital for cell growth, repair, and reproduction in organisms.

DNA Structure

DNA or deoxyribonucleic acid is a type of molecule known as a nucleic acid. It consists of a 5-carbon deoxyribose sugar, a phosphate, and a nitrogenous base. Double-stranded DNA consists of two spiral nucleic acid chains that are twisted into a double helix shape. This twisting allows DNA to be more compact. In order to fit within the nucleus, DNA is packed into tightly coiled structures called chromatin. Chromatin condenses to form chromosomes during cell division. Prior to DNA replication, the chromatin loosens giving cell replication machinery access to the DNA strands.

02
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Preparation For Replication

DNA Replication Fork
Computer artwork of a DNA (deoxyribonucleic acid) molecule during replication. DNA is composed of two strands. Each strand consists of a sugar-phosphate backbone (gray) attached to nucleotide bases. During replication the two strands unwind and separate, forming a replication bubble that enlarges to form a Y-shaped molecule termed a replication fork. It is here that daughter strands form a the parent DNA acts as a template for the construction of a new matching strand In this way the sequence of bases (or genetic information) along the DNA molecule is replicated. EQUINOX GRAPHICS/Science Photo Library/Getty Images

Step 1: Replication Fork Formation

Before DNA can be replicated, the double stranded molecule must be “unzipped” into two single strands. DNA has four bases called adenine (A), thymine (T), cytosine (C) and guanine (G) that form pairs between the two strands. Adenine only pairs with thymine and cytosine only binds with guanine. In order to unwind DNA, these interactions between base pairs must be broken. This is performed by an enzyme known as DNA helicase. DNA helicase disrupts the hydrogen bonding between base pairs to separate the strands into a Y shape known as the replication fork. This area will be the template for replication to begin.

DNA is directional in both strands, signified by a 5' and 3' end. This notation signifies which side group is attached the DNA backbone. The 5' end has a phosphate (P) group attached, while the 3' end has a hydroxyl (OH) group attached. This directionality is important for replication as it only progresses in the 5' to 3' direction. However, the replication fork is bi-directional; one strand is oriented in the 3' to 5' direction (leading strand) while the other is oriented 5' to 3' (lagging strand). The two sides are therefore replicated with two different processes to accommodate the directional difference.

Replication Begins

Step 2: Primer Binding

The leading strand is the simplest to replicate. Once the DNA strands have been separated, a short piece of RNA called a primer binds to the 3' end of the strand. The primer always binds as the starting point for replication. Primers are generated by the enzyme DNA primase.

 

03
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DNA Replication: Elongation

DNA Polymerases - Replication
DNA polymerases (blue) attach themselves to the DNA and elongate the new strands by adding nucleotide bases. BSIP/UIG/Getty Images

Step 3: Elongation

Enzymes known as DNA polymerases are responsible creating the new strand by a process called elongation. There are five different known types of DNA polymerases in bacteria and human cells. In bacteria such as E. coli, polymerase III is the main replication enzyme, while polymerase I, II, IV and V are responsible for error checking and repair. DNA polymerase III binds to the strand at the site of the primer and begins adding new base pairs complementary to the strand during replication. In eukaryotic cells, polymerases alpha, delta, and epsilon are the primary polymerases involved in DNA replication. Because replication proceeds in the 5' to 3' direction on the leading strand, the newly formed strand is continuous.

The lagging strand begins replication by binding with multiple primers. Each primer is only several bases apart. DNA polymerase then adds pieces of DNA, called Okazaki fragments, to the strand between primers. This process of replication is discontinuous as the newly created fragments are disjointed.

Step 4: Termination

Once both the continuous and discontinuous strands are formed, an enzyme called exonuclease removes all RNA primers from the original strands. These primers are then replaced with appropriate bases. Another exonuclease “proofreads” the newly formed DNA to check, remove and replace any errors. Another enzyme called DNA ligase joins Okazaki fragments together forming a single unified strand. The ends of the linear DNA present a problem as DNA polymerase can only add nucleotides in the 5′ to 3′ direction. The ends of the parent strands consist of repeated DNA sequences called telomeres. Telomeres act as protective caps at the end of chromosomes to prevent nearby chromosomes from fusing. A special type of DNA polymerase enzyme called telomerase catalyzes the synthesis of telomere sequences at the ends of the DNA. Once completed, the parent strand and its complementary DNA strand coils into the familiar double helix shape. In the end, replication produces two DNA molecules, each with one strand from the parent molecule and one new strand.

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Replication Enzymes

DNA Polymerase
This is an illustration of a DNA polymerase molecule. DNA polymerase is an enzyme that synthesizes DNA. Callista Image/Cultura/Getty Images

DNA replication would not occur without enzymes that catalyze various steps in the process. Enzymes that participate in the eukaryotic DNA replication process include:

  • DNA helicase - unwinds and separates double stranded DNA as it moves along the DNA. It forms the replication fork by breaking hydrogen bonds between nucleotide pairs in DNA.
  • DNA primase - a type of RNA polymerase that generates RNA primers. Primers are short RNA molecules that act as templates for the starting point of DNA replication.
  • DNA polymerases - synthesize new DNA molecules by adding nucleotides to leading and lagging DNA strands.
  • Topoisomerase or DNA Gyrase - unwinds and rewinds DNA strands to prevent the DNA from becoming tangled or supercoiled.
  • Exonucleases - group of enzymes that remove nucleotide bases from the end of a DNA chain.
  • DNA ligase - joins DNA fragments together by forming phosphodiester bonds between nucleotides.

 

05
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DNA Replication Summary

DNA Replication
Artwork illustrating a mechanism for replication of DNA, the substance of genetic inheritance. The DNA double helix consists of two strands of nucleotides (sugar, phosphate & base) lying antiparallel; each strand is joined through hydrogen bonding between the bases adenine (A), thymine (T), Guanine (G) and cytosine (C). A always pairs with T, and C with G; the resulting base sequence along the molecule determines genetic information. Conservation of information during cell replication occurs through each parent strand acting as a template for the formation of a complementary strand in a daughter molecule. Francis Leroy, BIOCOSMOS/Science Photo Library/Getty Images

DNA replication is the production of identical DNA helices from a single double-stranded DNA molecule. Each molecule consists of a strand from the original molecule and a newly formed strand. Prior to replication, the DNA uncoils and strands separate. A replication fork is formed which serves as a template for replication. Primers bind to the DNA and DNA polymerases add new nucleotide sequences in the 5′ to 3′ direction. This addition is continuous in the leading strand and fragmented in the lagging strand. Once elongation of the DNA strands is complete, the strands are checked for errors, repairs are made, and telomere sequences are added to the ends of the DNA.