Types
- Type I enzymes- The DNA at any distant position from a recognition site is cut by these restriction enzymes. The unique ATP and S-adenosyl-L-methionine are what trigger the activation of these restriction enzymes. These enzymes are still unique and possess multiple functions, including methylase and restriction digestion.
- Type II enzymes- This kind of restriction enzyme can cleave or split DNA from a location close to the actual recognition site. It does need a lot of magnesium to work properly. This enzyme is functional for single use and is independent of methylase.
- Type III enzymes- Additionally, this restriction enzyme fragments DNA at a location close to the real recognition site. It needs ATP to function, but it doesn’t need any hydrolase. The beginning of the reaction requires S-adenosyl-L-methionine. However, once the reaction starts, enzyme activity is irrelevant. With the aid of a modification methylase, this restriction enzyme can aid in the digestion of DNA.
- Type IV- The type IV restriction enzyme is a unique endonuclease that only works on DNA that has been altered. This restriction endonuclease, which works on DNA, is frequently employed in the biotechnology industry. Methylated and hydroxymethylated enzymes are two examples of restriction enzymes.
- Type V- The Type V restriction enzyme does not function as a DNA reaction enzyme. An RNA guide known as the gRNAs catalyzes this restriction endonuclease’s action on RNA.
Artificial Restriction Enzymes
By linking an artificial nuclease domain to a synthetic DNA-binding domain, restriction enzymes can be created. Such synthetic restriction enzymes can bind to specific DNA sequences and can target big DNA locations (up to 36 bp). The most popular artificial restriction enzymes are zinc finger nucleases, which are typically utilized in genetic engineering but can also be used in more conventional gene cloning procedures. The DNA-binding domain of TAL effectors serves as the foundation for additional synthetic restriction enzymes.
In 2013, the genome editing tool CRISPR-Cas9, based on a prokaryotic virus defense mechanism, was developed. It was quickly embraced in labs. Also created are synthetic ribonucleases that function as RNA restriction enzymes. A PNAzyme is a PNA-based system that mimics ribonucleases for a particular RNA sequence and cleaves at a non-base-paired region (RNA bulge) of the targeted RNA generated when the enzyme binds the RNA. This region is known as the RNA bulge. This enzyme exhibits selectivity by only cleaving at one of two potential cleavage sites that are either kinetically favored or do not have a mismatch.
Restriction Enzymes
Restriction enzyme is a bacterial protein that cleaves DNA at particular locations, these sites are called restricted sites. The restriction enzymes guard against bacteriophages in living bacteria. They identify the bacteriophage and cleave it at its restriction sites, destroying its DNA. Important genetic engineering tools include restriction enzymes. They may be separated from bacteria and applied in research facilities. The recognition sequences, or short and distinct nucleotide sequences, are recognized by restriction enzymes in DNA. When a DNA sequence is recognized by the restriction enzyme, it hydrolyzes the bond between neighboring nucleotides and cleaves the DNA molecule. The bacteria use the enzyme methylases to add the methyl group at the adenine or cytosine bases within the recognition sequence, preventing the DNA sequences from disintegrating.