What do endonucleases do

Accessed 13 Nov. More Definitions for endonuclease. Subscribe to America's largest dictionary and get thousands more definitions and advanced search—ad free! Log in Sign Up. Save Word. Definition of endonuclease. Adalja explains. First Known Use of endonuclease , in the meaning defined above. Learn More About endonuclease.

Time Traveler for endonuclease The first known use of endonuclease was in See more words from the same year. Statistics for endonuclease Look-up Popularity. In addition, there are intrinsic errors and unusual structures, which are formed during replication or recombination, and they must be corrected by the various repair protein machineries to avoid alterations of the base sequences or entanglement of the DNA.

These DNA repair proteins may function independently, but in many cases, they form complexes to perform more efficient repair reactions. In the repair complexes, nucleases play important roles in eliminating the damaged or mismatched nucleotides. They also recognize the replication or recombination intermediates to facilitate the following reaction steps through the cleavage of DNA strands Table 1. Nucleases can be regarded as molecular scissors, which cleave phosphodiester bonds between the sugars and the phosphate moieties of DNA.

They contain conserved minimal motifs, which usually consist of acidic and basic residues forming the active site. These active site residues coordinate catalytically essential divalent cations, such as magnesium, calcium, manganese or zinc, as a cofactor. However, the requirements for actual cleavage, such as the types and the numbers of metals, are very complicated, but are not common among the nucleases.

It appears that the major role of the metals is to stabilize intermediates, thereby facilitating the phosphoryl transfer reactions.

Cleavage reactions occur either at the end or within DNA, and thus DNA nucleases are categorized as exonucleases and endonucleases, respectively Figure 1. Schematic diagram of the nuclease activity. The two strands of DNA are schematically drawn. The cleavage made by the nuclease is represented by arrowhead. This review describes the three-dimensional 3D structural views of the actions of various nucleases involved in many DNA repair pathways. The rapidly accumulating genomic, biochemical and structural data have allowed us to classify various nucleases into folding families.

In general, the nucleases involved in DNA repair recognize the damaged moiety through the remarkably large deformation of DNA duplexes, and thus in terms of their DNA recognition mode, they apparently differ from the sequence-specific endonucleases, such as the restriction enzymes. The active sites of DNA repair nucleases have some similarity with other nucleases, including the metal-coordinating residues; however, they also display pronounced diversity.

Most DNA polymerases in prokaryotes and eukaryotes are composed of two different enzymes, a polymerase and an exonuclease, encoded within the same polypeptide, but sometimes they are formed by different subunits.

Deletion of these proofreading nucleases results in lethal or strong mutator phenotypes in bacteria Fijalkowska and Schaaper, and in yeast Morrison et al. Nuclease associated DNA repair pathways. The substrate DNAs are drawn schematically and the arrowheads denote nuclease cleavage. RNA regions are drawn in bold line. The removal of Okazaki fragments is another important process in replication.

Most of the Okazaki fragments are eliminated by RNaseH, enzyme ubiquitously present in all living organisms. In eukaryotes and in archaea, FEN1 endonucleases also participate in the removal of Okazaki fragments reviewed in Lieber, FEN1 is a multi-functional enzyme. This latter activity is required to eliminate non-homologous tails in base excision repair and in recombination intermediates. The replication process is stalled by various modes of DNA damage.

Upon the halt of fork progression, the DNA polymerase and other protein complexes abandon the replication fork. The remaining fork must be processed by various fork-specific protein machineries. Genetic and biochemical analyses have revealed that this endonuclease is completely conserved in eukaryotes, while its homolog has been found in archaea.

The loss of Mus81 in yeast causes UV or methylation damage sensitivity Interthal and Heyer, and defects in sporulation Mullen et al.

Abasic sites within DNA duplexes are frequently produced by the actions of various DNA glycosylases involved in the base excision repair pathway, in addition to the spontaneous hydrolysis of bases.

These apyrimidine or apurine AP sites are removed by AP endonucleases which cleave the phosphdiester bond next to an abasic site Figure 2 reviewed in Mol et al. Interestingly, these two enzymes show no sequence similarity to each other; although their AP endonuclease activities are quite similar. In eukaryotes, there seems to be a single, major AP endonuclease working in each organism. APN1, the yeast homolog of E. The absence of APN1 results in enhanced sensitivity to oxidative damage and alkylating agents Ramotar et al.

Mammalian organisms, including humans, bear Ape1, which shares sequence similarity with E. In addition to the AP endonuclease activity, Ape1 also plays a major role in sensing the redox state of the cell Xanthoudakis et al. The loss of Ape1 generates embryonic lethality in mice Wilson and Thompson, In prokaryotes, mismatch repair is conducted mainly by the MutSLH proteins, while the Vsr protein is responsible for mismatches in certain sequences reviewed in Modrich and Lahue, ; Yang, ; Tsutakawa and Morikawa, The cleavage activity of MutH is enhanced by the MutL protein, although its mechanism remains unclear.

Vsr makes an incision next to the mismatched base. These forms of damage involve those generated by the UV radiation and the large adducts produced by various chemicals. In bacteria, this dual incision is performed by the UvrB-UvrC complex. Deletions or mutations introduced into these nucleases cause sensitivity to UV damage, and result in cancer formation. In addition, abnormalities of these proteins cause defects in neural development. Double strand breaks are generated by the accidental halt of fork progression during replication or by ionizing radiation and strand incision chemicals.

They are also generated as an intermediate state during meiosis and V D J recombination. These double strand breaks are repaired through the two main pathways of non-homologous end joining and homologous recombination.

In either case, the ends of the double strand breaks must be processed to initiate the repair reaction Figure 2. Mre11 is a multi-functional nuclease involved in the processing of the DNA ends or hairpin structures reviewed in D'Amours and Jackson, While Mre11 itself exhibits a ssDNA exonuclease activity, its complex with Rad50 processes double strand break ends.

Mutations introduced into Mre11 cause an ataxia-telangiectasia-like disorder Stewart et al. V D J recombination involves a reaction process, in which hairpin DNAs are opened, and subsequently, both ends are connected. Although Artemis alone possesses a ssDNA exonuclease activity, its complex formation with DNA-PK allows the processing of the double strand break ends to open the hairpin structure.

Defects in each protein cause severe immunodeficiency Blunt et al. In homologous recombination, two homologous DNA strands are paired and are connected by D-loop structures or Holliday junction intermediates. In bacteria, the RuvC protein cleaves the Holliday junction at two symmetrical sites near the junction center to resolve the junction into two dsDNAs Figure 2.

Similar junction resolving enzymes have also been found in other bacteria, bacteriophages, and archaea reviewed in Sharples, The primary sequences of nucleases are often poorly conserved, except for the motifs related to catalytic sites. The functional and biochemical properties of many nucleases have been studied extensively. However, in some cases, it is very difficult to identify the actual functional targets of the nucleases, because of their broad substrate specificity.

Nevertheless, many candidates for nucleases are available from various genome sequences, and their functional properties can be inferred by sequence comparisons with other well-studied nucleases.

For instance, Koonin and his associates have successfully classified nucleases, phosphoesterases, and phosphatases into several families, based on extensive data base analyses of the primary sequences Aravind and Koonin, a , b ; Aravind et al. This classification has also revealed the relationships between nucleases and identified several new nuclease families. In addition to the classifications of primary sequences, 3D structural data have been rapidly accumulating with respect to the proteins involved in DNA repair, including nucleases.

The classification of nucleases in terms of their 3D structures provides more defined properties, since it is accepted that the 3D structures are much less diverged and more closely related to the functions than the primary sequences. As a matter of fact, in the type II restriction endonucleases, all of the structures share the common core motif, which includes the active sites, and thus could be grouped into a single folding family, despite its primary sequence diversity reviewed in Pingoud and Jeltsch, Folding patterns of DNA repair nucleases.

The core folding is drawn schematically. The positions of the bound metals are marked by black circles. Representative repair nuclease of the folding is written in parenthesis. The RNaseH-like fold, which is one of the most ubiquitous architectures in the protein world, has been found in RuvC, RNaseH, integrase, transposase, and proofreading exonucleases Figure 3a.

The strand order is , with strand 2 anti-parallel to the others. The active site residues, which are constituted according to the DDE motif, are located on one side of the sheet. These three or sometimes four acidic residues coordinate the metals, which are essential for the catalytic reaction. The cocrystal structure with TMP revealed that the phosphate moiety is directly coordinated between the two metals, as it mimics the product DNA. However, it possesses a different strand order, which is defined as with strand 5 anti-parallel to the others.

The structures of restriction endonucleases revealed that their catalytic domains share common fold architecture Figure 3c. The strand order is , with strand 2, and in some cases, strand 5, anti-parallel to the others. Similar sequences are also found in several DNA repair nucleases, such as MutH, Hjc, and T7 endoI, which are categorized into essentially the same folding family. The active sites in endonucleases with the restriction endonuclease-like fold coordinate up to three metals depending upon the enzyme.

This fold was recently identified by the determination of the RecJ nuclease structure Yamagata et al. Previous sequence analyses have shown that this family includes RecJ and the phosphoesterases, which contain conserved phosphoesterase motifs Aravind and Koonin, a.

These residues, which constitute part of the active site, are likely to participate in the cleavage reaction. The cocrystal structure of Mre11 with Mn and dAMP shows two manganese ions bound to the active site, and these two metals are simultaneously coordinated to the phosphate moiety, thus mimicking the product-bound state Hopfner et al. It is also observed in some phosphatases, such as inositol 5-phosphatase. The crystal structures of DNaseI Suck et al. On the other hand, one Gorman et al.

They are categorized based on their mechanism of action. They are regularly used in genetic engineering to create recombinant DNA, which can be introduced into different cells of bacterial, plant or animal origin.

They can also be used in synthetic biology. It is a protein which plays an important role in the immunological defense of certain bacteria against DNA viruses.

It has become more well-known due to its uses in genetic engineering. Endonuclease Structure Type I and Type II restriction endonucleases are multisubunit complexes that include endonucleases and methylase activities. Type I restriction enzymes are capable of cleaving a random sites of approximately 1, base pairs from the recognition sequence.

There are two AP endonucleases in E. Mutations can also occur in endonucleases.