Endonuclease
Encyclopedia
Endonucleases are enzyme
s that cleave the phosphodiester bond
within a polynucleotide
chain, in contrast to exonuclease
s, which cleave phosphodiester bond
s at the end of a polynucleotide chain. Typically, a restriction site will be a palindromic sequence four to six nucleotides long. Most restriction endonucleases cleave the DNA strand unevenly, leaving complementary single-stranded ends. These ends can reconnect through hybridization and are termed "sticky ends." Once paired, the phosphodiester bonds of the fragments can be joined by DNA ligase. There are hundreds of restriction endonucleases known, each attacking a different restriction site. A given sample of DNA is likely to contain a recognition sequence for any restriction endonuclease. The DNA fragments cleaved by the same endonuclease can be joined together regardless of the origin of the DNA. Such DNA is called recombinant DNA; it has been artificially recombined. Restriction endonucleases (restriction enzyme
s) and are divided into three categories, Type I, Type II, and Type III, according to their mechanism of action. These enzymes are often used in genetic engineering
to make recombinant DNA
for introduction into bacterial, plant, or animal cells, as well as in synthetic biology
.
The commonly used notation for restriction endonucleases is of the form "vwxyz", where "vwx" names the life form (bacteria) where this restriction endonuclease may be found, "y" names the strain (and is optional), and "z" (in Roman numerals) indicates different restriction endonucleases in the same life form (bacteria). Thus for example, "EcoRI" means that the restriction endonuclease is found in Escherichia coli ("Eco"); strain RY13 ("R"), restriction endonuclease number "I". Another example: "HaeII" and "HaeIII" refer to bacterium Haemophilus aegyptius, number II and number III, respectively.
Restriction endonucleases come in several types. A restriction endonuclease typically requires a recognition site and a cleavage pattern (typically of nucleotide bases: A, C, G, T). If the recognition site is outside the region of the cleavage pattern, then the restriction endonuclease is referred to as Type I. If the recognition sequence overlaps with the cleavage sequence, then the restriction endonuclease restriction enzyme
is Type II.
Restriction endonucleases may be found that cleave standard dsDNA (double-stranded DNA), or ssDNA (single-stranded DNA), or even RNA. This discussion is restricted to dsDNA, however, the discussion can be extended:
In addition, research is now underway to construct synthetic or artificial restriction endonucleases, especially with recognition sites that are unique within a genome.
Restriction endonucleases or restriction enzymes typically cleave in two ways: blunt-ended or sticky-ended patterns. An example of a Type I restriction endonuclease, see
ENases are widespread amongst prokaryotes (bacteria and archaea), but are produced by some eukaryotic organisms as well, although this is very rare.
Some endonucleases have actions on RNA, such as the Dicer
enzyme, which initiates the formation of RNA-induced silencing complexes. These may also be termed endoribonuclease
s.
Enzyme
Enzymes are proteins that catalyze chemical reactions. In enzymatic reactions, the molecules at the beginning of the process, called substrates, are converted into different molecules, called products. Almost all chemical reactions in a biological cell need enzymes in order to occur at rates...
s that cleave the phosphodiester bond
Phosphodiester bond
A phosphodiester bond is a group of strong covalent bonds between a phosphate group and two 5-carbon ring carbohydrates over two ester bonds. Phosphodiester bonds are central to all known life, as they make up the backbone of each helical strand of DNA...
within a polynucleotide
Polynucleotide
A polynucleotide molecule is a biopolymer composed of 13 or more nucleotide monomers covalently bonded in a chain. DNA and RNA are examples of polynucleotides with distinct biological function. The prefix poly comes from the ancient Greek πολυς...
chain, in contrast to exonuclease
Exonuclease
Exonucleases are enzymes that work by cleaving nucleotides one at a time from the end of a polynucleotide chain. A hydrolyzing reaction that breaks phosphodiester bonds at either the 3’ or the 5’ end occurs. Its close relative is the endonuclease, which cleaves phosphodiester bonds in the middle ...
s, which cleave phosphodiester bond
Phosphodiester bond
A phosphodiester bond is a group of strong covalent bonds between a phosphate group and two 5-carbon ring carbohydrates over two ester bonds. Phosphodiester bonds are central to all known life, as they make up the backbone of each helical strand of DNA...
s at the end of a polynucleotide chain. Typically, a restriction site will be a palindromic sequence four to six nucleotides long. Most restriction endonucleases cleave the DNA strand unevenly, leaving complementary single-stranded ends. These ends can reconnect through hybridization and are termed "sticky ends." Once paired, the phosphodiester bonds of the fragments can be joined by DNA ligase. There are hundreds of restriction endonucleases known, each attacking a different restriction site. A given sample of DNA is likely to contain a recognition sequence for any restriction endonuclease. The DNA fragments cleaved by the same endonuclease can be joined together regardless of the origin of the DNA. Such DNA is called recombinant DNA; it has been artificially recombined. Restriction endonucleases (restriction enzyme
Restriction enzyme
A Restriction Enzyme is an enzyme that cuts double-stranded DNA at specific recognition nucleotide sequences known as restriction sites. Such enzymes, found in bacteria and archaea, are thought to have evolved to provide a defense mechanism against invading viruses...
s) and are divided into three categories, Type I, Type II, and Type III, according to their mechanism of action. These enzymes are often used in genetic engineering
Genetic engineering
Genetic engineering, also called genetic modification, is the direct human manipulation of an organism's genome using modern DNA technology. It involves the introduction of foreign DNA or synthetic genes into the organism of interest...
to make recombinant DNA
Recombinant DNA
Recombinant DNA molecules are DNA sequences that result from the use of laboratory methods to bring together genetic material from multiple sources, creating sequences that would not otherwise be found in biological organisms...
for introduction into bacterial, plant, or animal cells, as well as in synthetic biology
Synthetic biology
Synthetic biology is a new area of biological research that combines science and engineering. It encompasses a variety of different approaches, methodologies, and disciplines with a variety of definitions...
.
The commonly used notation for restriction endonucleases is of the form "vwxyz", where "vwx" names the life form (bacteria) where this restriction endonuclease may be found, "y" names the strain (and is optional), and "z" (in Roman numerals) indicates different restriction endonucleases in the same life form (bacteria). Thus for example, "EcoRI" means that the restriction endonuclease is found in Escherichia coli ("Eco"); strain RY13 ("R"), restriction endonuclease number "I". Another example: "HaeII" and "HaeIII" refer to bacterium Haemophilus aegyptius, number II and number III, respectively.
Restriction endonucleases come in several types. A restriction endonuclease typically requires a recognition site and a cleavage pattern (typically of nucleotide bases: A, C, G, T). If the recognition site is outside the region of the cleavage pattern, then the restriction endonuclease is referred to as Type I. If the recognition sequence overlaps with the cleavage sequence, then the restriction endonuclease restriction enzyme
Restriction enzyme
A Restriction Enzyme is an enzyme that cuts double-stranded DNA at specific recognition nucleotide sequences known as restriction sites. Such enzymes, found in bacteria and archaea, are thought to have evolved to provide a defense mechanism against invading viruses...
is Type II.
Restriction endonucleases may be found that cleave standard dsDNA (double-stranded DNA), or ssDNA (single-stranded DNA), or even RNA. This discussion is restricted to dsDNA, however, the discussion can be extended:
- Standard dsDNA
- Non-standard DNA
- Holliday junctions Holliday junctionHolliday junctionA Holliday junction is a mobile junction between four strands of DNA. The structure is named after Robin Holliday, who proposed it in 1964 to account for a particular type of exchange of genetic information he observed in yeast known as homologous recombination...
- Triple-stranded DNA triple-stranded DNATriple-stranded DNAA triple-stranded DNA is a structure of DNA in which three oligonucleotides wind around each other and form a triple helix. In this structure, one strand binds to a B-form DNA double helix through Hoogsteen or reversed Hoogsteen hydrogen bonds....
, quadruple-stranded DNA (G-quadruplexG-quadruplexIn molecular biology, G-quadruplexes are nucleic acid sequences that are rich in guanine and are capable of forming a four-stranded structure...
), etc. - Double-stranded hybrids of DNA and RNA (one strand is DNA, the other strand is RNA)
- Synthetic or artificial DNA (for example, containing bases other than A, C, G, T, refer to the work of Eric T. Kool). Research with synthetic codons, refer to the research by S. Benner, and enlarging the amino acid set in polypeptides, thus enlarging the proteome or proteomicsProteomicsProteomics is the large-scale study of proteins, particularly their structures and functions. Proteins are vital parts of living organisms, as they are the main components of the physiological metabolic pathways of cells. The term "proteomics" was first coined in 1997 to make an analogy with...
, see the research by P. Schultz.
In addition, research is now underway to construct synthetic or artificial restriction endonucleases, especially with recognition sites that are unique within a genome.
Restriction endonucleases or restriction enzymes typically cleave in two ways: blunt-ended or sticky-ended patterns. An example of a Type I restriction endonuclease, see
Common endonucleases
Restriction endonucleases (ENases) are most commonly produced by bacteria, and can be used to map a piece of DNA.ENases are widespread amongst prokaryotes (bacteria and archaea), but are produced by some eukaryotic organisms as well, although this is very rare.
Some endonucleases have actions on RNA, such as the Dicer
Dicer
Dicer is an endoribonuclease in the RNase III family that cleaves double-stranded RNA and pre-microRNA into short double-stranded RNA fragments called small interfering RNA about 20-25 nucleotides long, usually with a two-base overhang on the 3' end...
enzyme, which initiates the formation of RNA-induced silencing complexes. These may also be termed endoribonuclease
Endoribonuclease
A Endoribonuclease is a ribonuclease endonuclease. It cleaves either single-stranded or double-stranded RNA, depending on the enzyme. Example includes both single proteins like RNase III, RNase A, RNase T1, and RNase H, and also complexes of proteins like RNase P and the RNA-induced silencing...
s.