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A nick is a discontinuity in a double stranded
DNA Deoxyribonucleic acid (; DNA) is a polymer composed of two polynucleotide chains that coil around each other to form a double helix. The polymer carries genetic instructions for the development, functioning, growth and reproduction of al ...
molecule where there is no
phosphodiester bond In chemistry, a phosphodiester bond occurs when exactly two of the hydroxyl groups () in phosphoric acid react with hydroxyl groups on other molecules to form two ester bonds. The "bond" involves this linkage . Discussion of phosphodiesters is d ...
between adjacent
nucleotide Nucleotides are Organic compound, organic molecules composed of a nitrogenous base, a pentose sugar and a phosphate. They serve as monomeric units of the nucleic acid polymers – deoxyribonucleic acid (DNA) and ribonucleic acid (RNA), both o ...
s of one
strand Strand or The Strand may refer to: Topography *The flat area of land bordering a body of water, a: ** Beach ** Shoreline * Strand swamp, a type of swamp habitat in Florida Places Africa * Strand, Western Cape, a seaside town in South Africa * ...
. They typically occur through damage or
enzyme An enzyme () is a protein that acts as a biological catalyst by accelerating chemical reactions. The molecules upon which enzymes may act are called substrate (chemistry), substrates, and the enzyme converts the substrates into different mol ...
action. Nicks allow DNA strands to untwist during replication, and are also thought to play a role in the
DNA mismatch repair DNA mismatch repair (MMR) is a system for recognizing and repairing erroneous insertion, deletion, and mis-incorporation of nucleobase, bases that can arise during DNA replication and Genetic recombination, recombination, as well as DNA repair, ...
mechanisms that fix errors on both the leading and lagging daughter strands.


Formation of nicks

The diagram shows the effects of nicks on intersecting DNA in a twisted
plasmid A plasmid is a small, extrachromosomal DNA molecule within a cell that is physically separated from chromosomal DNA and can replicate independently. They are most commonly found as small circular, double-stranded DNA molecules in bacteria and ...
. Nicking can be used to dissipate the energy held up by intersecting states. The nicks allow the DNA to take on a circular shape. Nicked DNA can be the result of DNA damage or purposeful, regulated biomolecular reactions carried out in the cell. During processing, DNA can be nicked by physical shearing, over-drying, or enzymes. Excessive rough handling in pipetting or vortexing creates physical stress that can lead to breaks and nicks in DNA. Overdrying of DNA can also break the phosphodiester bond in DNA and result in nicks. Nicking enzymes can assist with this process. A nick in DNA can be formed by the hydrolysis and subsequent removal of a phosphate group within the helical backbone. This leads to a different DNA conformation, where a hydrogen bond forms in place of the missing piece of the DNA backbone in order to preserve the structure.


Repair of nicks

Ligase In biochemistry, a ligase is an enzyme that can catalyze the joining ( ligation) of two molecules by forming a new chemical bond. This is typically via hydrolysis of a small pendant chemical group on one of the molecules, typically resulting i ...
s are versatile and ubiquitous
enzyme An enzyme () is a protein that acts as a biological catalyst by accelerating chemical reactions. The molecules upon which enzymes may act are called substrate (chemistry), substrates, and the enzyme converts the substrates into different mol ...
s that join the 3’ hydroxyl and 5’ phosphate ends to form a phosphodiester bond, making them essential in nicked DNA repair, and ultimately genome fidelity. This biological role has also been extremely valuable in sealing the
sticky ends DNA ends refer to the properties of the ends of linear DNA molecules, which in molecular biology are described as "sticky" or "blunt" based on the shape of the complementary strands at the terminus. In sticky ends, one strand is longer than the o ...
of plasmids in molecular cloning. Their importance is attested by the fact most organisms have multiple ligases dedicated to specific pathways of repairing DNA. In eubacteria, these ligases are powered by
NAD+ Nicotinamide adenine dinucleotide (NAD) is a coenzyme central to metabolism. Found in all living cells, NAD is called a dinucleotide because it consists of two nucleotides joined through their phosphate groups. One nucleotide contains an ade ...
rather than ATP. Each nick site requires 1 ATP or 1 NAD+ to power the ligase repair. In order to join these fragments, the ligase progresses through three steps: # Addition of an
adenosine monophosphate Adenosine monophosphate (AMP), also known as 5'-adenylic acid, is a nucleotide. AMP consists of a phosphate group, the sugar ribose, and the nucleobase adenine. It is an ester of phosphoric acid and the nucleoside adenosine. As a substituent it t ...
(AMP) group to the enzyme, referred to as adenylylation # Adenosine monophosphate transfer to the DNA # Nick sealing, or phosphodiester bond formation. One particular example of a ligase catalyzing nick closure is the ''E. coli'' NAD+ dependent DNA ligase, LigA. LigA is a relevant example as it is structurally similar to a clade of enzymes found across all types of bacteria. Ligases have a metal binding site which is capable of recognizing nicks in DNA. The ligase forms a DNA-adenylate complex, assisting recognition. With human DNA ligase, this forms a crystallized complex. The complex, which has a DNA–adenylate intermediate, allows DNA ligase I to institute a conformational change in the DNA for the isolation and subsequent repair of the DNA nick.


Biological implications


Role in mismatch repair

Single-stranded nicks act as recognizable markers to help the repair machinery distinguish the newly synthesized strand (daughter strand) from the template strand (parental strand). DNA mismatch repair (MMR) is an important DNA repair system that helps maintain genome plasticity by correcting mismatches, or non Watson-Crick base pairs in a DNA duplex. Some sources of mismatched base pairs include replication errors and
deamination Deamination is the removal of an amino group from a molecule. Enzymes that catalysis, catalyse this reaction are called deaminases. In the human body, deamination takes place primarily in the liver; however, it can also occur in the kidney. In s ...
of 5-methylcytosine DNA to form thymine. MMR in most bacteria and
eukaryote The eukaryotes ( ) constitute the Domain (biology), domain of Eukaryota or Eukarya, organisms whose Cell (biology), cells have a membrane-bound cell nucleus, nucleus. All animals, plants, Fungus, fungi, seaweeds, and many unicellular organisms ...
s is directed to the erroneous strand of the mismatched duplex through recognition of strand discontinuities, while MMR in ''E. coli'' and closely related bacteria is directed to the strand on the basis of the absence of
methylation Methylation, in the chemistry, chemical sciences, is the addition of a methyl group on a substrate (chemistry), substrate, or the substitution of an atom (or group) by a methyl group. Methylation is a form of alkylation, with a methyl group replac ...
. Nicking endonucleases introduce strand discontinuities, or DNA nicks, for both respective systems. Mut L homologues from eukaryotes and most bacteria incise the discontinuous strand to introduce the entry or termination point for the excision reaction. Similarly, in ''E. coli'', Mut H nicks the unmethylated strand of the duplex to introduce the entry point of excision. For eukaryotes specifically, the mechanism of DNA replication elongation between the leading and lagging strand differs. On the lagging strand, nicks exist between
Okazaki fragments Okazaki fragments are short sequences of DNA nucleotides (approximately 150 to 200 base pairs long in eukaryotes) which are synthesized discontinuously and later linked together by the enzyme DNA ligase to create the lagging strand during DN ...
and are easily recognizable by the DNA mismatch repair machinery prior to ligation. Due to the continuous replication that occurs on the leading strand, the mechanism there is slightly more complex. During replication,
ribonucleotide In biochemistry, a ribonucleotide is a nucleotide containing ribose as its pentose component. It is considered a molecular precursor of nucleic acids. Nucleotides are the basic building blocks of DNA and RNA. Ribonucleotides themselves are basic mo ...
s are added by replication enzymes and these ribonucleotides are nicked by an enzyme called RNase H2. Together, the presence of a nick and a ribonucleotide make the leading strand easily recognizable to the DNA mismatch repair machinery.
Nick translation Nick translation (or head translation), developed in 1977 by Peter Rigby and Paul Berg, is a tagging technique in molecular biology in which DNA polymerase I is used to replace some of the nucleotides of a DNA sequence with their labeled analogue ...
is a biological process in which a single-stranded DNA nick serves as the marker for
DNA polymerase A DNA polymerase is a member of a family of enzymes that catalyze the synthesis of DNA molecules from nucleoside triphosphates, the molecular precursors of DNA. These enzymes are essential for DNA replication and usually work in groups to create t ...
to excise and replace possibly damaged nucleotides. At the end of the segment that DNA polymerase acts on,
DNA ligase DNA ligase is a type of enzyme that facilitates the joining of DNA strands together by catalyzing the formation of a phosphodiester bond. It plays a role in repairing single-strand breaks in duplex DNA in living organisms, but some forms (such ...
must repair the final segment of the DNA backbone in order to complete the repair process. In a lab setting, this can be used to introduce
fluorescent Fluorescence is one of two kinds of photoluminescence, the emission of light by a substance that has absorbed light or other electromagnetic radiation. When exposed to ultraviolet radiation, many substances will glow (fluoresce) with color ...
or other tagged nucleotides by purposefully inducing site-specific, single-stranded nicks in DNA
in vitro ''In vitro'' (meaning ''in glass'', or ''in the glass'') Research, studies are performed with Cell (biology), cells or biological molecules outside their normal biological context. Colloquially called "test-tube experiments", these studies in ...
and then adding the nicked DNA to an environment rich in DNA polymerase and tagged nucleotide. The DNA polymerase then replaces the DNA nucleotides with the tagged ones, starting at the site of the single-stranded nick.


Role in replication and transcription

Nicked DNA plays an important role in many biological functions. For instance, single-stranded nicks in DNA may serve as purposeful biological markers for the enzyme
topoisomerase DNA topoisomerases (or topoisomerases) are enzymes that catalyze changes in the topological state of DNA, interconverting relaxed and supercoiled forms, linked (catenated) and unlinked species, and knotted and unknotted DNA. Topological issues in ...
that unwinds packed DNA and is critical to
DNA replication In molecular biology, DNA replication is the biological process of producing two identical replicas of DNA from one original DNA molecule. DNA replication occurs in all life, living organisms, acting as the most essential part of heredity, biolog ...
and transcription. In these instances, nicked DNA is not the result of unwanted cell damage. Topoisomerase-1 preferentially acts at nicks in DNA to cleave adjacent to the nick and then winds or unwinds the complex topologies associated with packed DNA. Here, the nick in the DNA serves as a marker for single strand breakage and subsequent unwinding. It is possible that this is not a highly conserved process. Topoisomerase may cause short deletions when it cleaves bonds, because both full-length DNA products and short deletion strands are seen as products of topoisomerase cleavage while inactive mutants only produced full-length DNA strands. Nicks in DNA also give rise to different structural properties, can be involved in repairing damages caused by
ultraviolet Ultraviolet radiation, also known as simply UV, is electromagnetic radiation of wavelengths of 10–400 nanometers, shorter than that of visible light, but longer than X-rays. UV radiation is present in sunlight and constitutes about 10% of ...
radiation, and are used in the primary steps that allow for
genetic recombination Genetic recombination (also known as genetic reshuffling) is the exchange of genetic material between different organisms which leads to production of offspring with combinations of traits that differ from those found in either parent. In eukaryot ...
. Nick idling is a biological process in which DNA polymerase may slow or stop its activity of adding bases to a new daughter strand during DNA replication at a nick site. This is particularly relevant to Okazaki fragments in lagging strand in double stranded DNA replication because the direction of replication is opposite to the direction of DNA polymerase, therefore nick idling plays a role in stalling the complex as it replicates in the reverse direction in small fragments (Okazaki fragments) and has to stop and reposition itself in between each and every fragment length of DNA. DNA structure changes when a single-stranded nick is introduced. Stability is decreased as a break in the phosphodiester backbone allows DNA to unwind, as the built up stress from twisting and packing is not being resisted as strongly anymore. Nicked DNA is more susceptible to degradation due to this reduced stability.


In bacteria

The ''nic'' site or nick region is found within the origin of transfer (''oriT'') site and is a key in starting
bacterial conjugation Bacterial conjugation is the transfer of genetic material between Bacteria, bacterial cells by direct cell-to-cell contact or by a bridge-like connection between two cells. This takes place through a pilus. It is a parasexual cycle, parasexual mode ...
. A single strand of DNA, called the T-strand, is cut at ''nic'' by an enzyme called relaxase. This single strand is eventually transferred to the recipient cell during the process of bacterial conjugation. Before this cleavage can occur, however, it is necessary for a group of proteins to attach to the ''oriT'' site. This group of proteins is called the relaxosome. It is thought that portions of the ''oriT'' site are bent in a way that creates interaction between the relaxosome proteins and the ''nic'' site. Cleaving the T-strand involves relaxase cutting a phosphodiester bond at the nic site. The cleaved strand is left with a hydroxyl group at the 3' end, which may allow for the strand to form a circular plasmid after moving into the recipient cell.


Role in meiosis

DNA nicks promote crossover formation during
meiosis Meiosis () is a special type of cell division of germ cells in sexually-reproducing organisms that produces the gametes, the sperm or egg cells. It involves two rounds of division that ultimately result in four cells, each with only one c ...
, and such nicks are protected from ligation by Exonuclease 1 (Exo1).


References

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