Endonucleases

Endonucleases are enzymes that cleave the phosphodiester bond within a polynucleotide chain. Some, such as Deoxyribonuclease I, cut DNA relatively nonspecifically (without regard to sequence), while many, typically called restriction endonucleases or restriction enzymes, cleave only at very specific nucleotide sequences.

Restriction enzymes are endonucleases from eubacteria and archaea that recognize a specific DNA sequence. The nucleotide sequence recognized for cleavage by a restriction enzyme is called the restriction site. Typically, a restriction site will be a palindromic sequence about 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. 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; DNA formed by the joining of genes into new combinations.Restriction endonucleases (restriction enzymes) 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.

Ultimately, there are three categories of restriction endonucleases that relatively contribute to the cleavage of specific sequences. The types I and III are large multisubunit complexes that include both the endonucleases and methylase activities. Type I can cleave at random sites of about 1000 base pairs or more from the recognition sequence and it requires ATP as source of energy. Type II behaves slightly differently and was first isolated by Hamilton Smith in 1970. They are simpler versions of the endonucleases and require no ATP in their degradation processes. Some examples of type II restriction endonucleases include BamHI, EcoRI, EcoRV, and Haelll. Type III, however, cleaves the DNA at about 25 base pairs from the recognition sequence and also requires ATP in the process.

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