Restriction Endonucleases
Restriction Endonucleases
Nucleases are enzymes that break down nucleic acids (DNA or RNA) by cleaving the phosphodiester bonds between nucleotides. They can be classified based on the type of nucleic acid they act on and the way they cut it.
Restriction enzymes (or restriction endonucleases) are proteins that act as molecular scissors to cut DNA at specific sequences. They are found in bacteria and archaea, where they serve as a defense mechanism against viruses by cleaving foreign DNA (such as viral DNA) that enters the cell. These enzymes recognize specific, short DNA sequences, known as recognition sites (usually 4-8 base pairs long), and cut the DNA at or near those sequences.
There are two main types of restriction enzymes:
Type II restriction enzymes: These are the most commonly used in molecular biology for genetic manipulation. They cut the DNA at a defined location within their recognition site, producing either blunt ends or sticky ends (overhanging single-stranded regions).
Example: EcoRI, which recognizes the sequence GAATTC and cuts between G and A, creating sticky ends.
Type I and Type III restriction enzymes: These enzymes also cleave DNA, but their cutting sites are more complex and require ATP or other cofactors to function. These are less commonly used than Type II enzymes.
Applications:
DNA cloning: To cut DNA at specific locations for insertion into plasmids or vectors.
Restriction fragment length polymorphism (RFLP): Used for DNA fingerprinting or genetic mapping.
Gene editing: To prepare DNA fragments for experiments like gene knockout or insertion.