Nucleasesare enzymes that hydrolyze the phosphodiester bond in the backbone of nucleicacids. These nuclease enzymes play an important role in degradation of old and brokennucleic acids. There are two distinct types of nucleases: exonuclease andendonuclease. Exonuclease are enzymes that cleave a terminal nucleotide off anucleic acid.
Contrarily, endonuclease enzymes cleave a phosphodiester bondwithin a nucleic acid. Furthermore, endonuclease enzymes can be specific ornonspecific. Non-specific endonuclease cleaves at any internal bond or cleavesa phosphodiester bond at 3' carbon. Likewise, a specific endonuclease cleavesat a DNA sequence. Thus, endonucleases with extreme specificity arecalled restriction enzymesRestrictionenzymes recognize a specific sequence of DNA, normally the sequence is betweenfour to six base-pair long. There are many different types of restrictionenzymes such as type I, II, and III.
The article "Type II restrictionendonucleases—a historical perspective and more", introduces type IIrestriction enzymes. Type II restriction enzymes mainlyoccur in prokaryotes and are used to protect cells from viruses and otherinfectious DNA molecules. Type II restriction enzymes include EcoRI andEcoRV. EcoRV is a type II restriction enzyme derivedfrom Escherichia Coli. One concept that we did notcover in class was the different forms of restriction enzymes.
EcoRV is anenzyme which cleaves at every 5' GATATC 3' DNAsequence. The enzyme binds to DNA very loosely and scans for the 5' GATATC 3'sequence, once the sequence is found, EcoRV kinks DNA at an angle and cleavesat the 5' GATATC 3' sequence. EcoRV is the only form of restriction enzyme whichcauses a conformational change to occur in the DNA.Nonetheless,one form of type II restriction enzyme that we have covered in class is EcoRI,which is isolated from Escherichia Coli. The enzyme cleaves DNA at every 5' GTTAAC 3' DNA sequence. For instance, when DNA isdigested with RcoRI, it will cut DNA at every 5' GTTAAC 3' sequence.
Essentially,every person has a different TTAA sequence. Thus, everyone will have differentDNA fragments when digested with EcoRI. This form of DNA analysis isfundamental in paternity testing and criminal investigation. As previouslyestablished, we all have different TTAA fragments, thus during paternitytesting and criminal investigation you can easily identify who themother/father is of the child in question or who the criminal is when TTAAfragments are compared.Thediscovery of nucleases has advanced molecular biology, genetics, andbiotechnology. Nucleases are classified as enzymes which help cleave a DNAsequence.
There are two fundamental forms of nucleases: exonuclease andendonuclease. Restriction enzymes are a type of endonucleases. Inherently,there are different types of restriction enzymes. Two most important type IIrestriction enzymes are classified as EcoRI and EcoRV. Type II restrictionenzymes are primarily used to protect cells from viruses and other infectiousmolecules. Additionally, EcoRI is used in DNA analysis during paternity testingand criminal investigation.
Therefore, the discovery of restriction enzymesallows genes to be cloned, cleaved and overexpressed.
