Question

How does endonuclease function?

Answer 1

Restriction endonucleases are enzymes used to cut DNA at specific sites called restriction sites. Every restriction enzyme has a specific restriction site at which it cuts a DNA molecule. Restriction sequence for BamHI is GAATTC. The most abundantly used restriction enzymes are type II restriction enzymes because they cut at specific restriction sites only.

Complete answer:
These are divided into three groups.
Type I Restriction Endonucleases: They have a restriction sequence of 15 bp. DNA is cleaved 1000bp away from the 5' end of the sequence “TCA” located within the recognition site. Ex: EcoK-12, EcoB, etc. They methylate the DNA sequence at the site of recognition.

Type II Restriction Endonucleases: They are remarkably stable and induce cleavage either, in most cases, within or outside their recognition sequences,which are symmetrical. Two different types of cuts are generated depending on whether they cut both strands at the centre of the recognition sequence: The former cut will generate “blunt ends” with no nucleotide overhangs. The latter generates “sticky” or “cohesive” ends.

Type III Restriction Endonucleases: They cleave DNA in the immediate vicinity of their recognition sites, Ex;- EcoP1, EcoP15, Hind III etc. DNA is cut upto 20-30 base pairs away from the recognition site.
Working of Endonucleases: They recognize a specific sequence of nucleotides within the polynucleotide chain and cleave the phosphodiester bond , thus producing a double stranded cut in the polynucleotide chain. These cute products produce two types of ends:
Blunt ends: These blunt ended fragments can be Joined to any other DNA fragment with blunt ends.
Sticky ends: DNA fragments with complementary sticky ends can be combined to create new molecules which allows the creation and manipulation of DNA sequences from different sources.

Note:
Enzymes are believed to be evolved by bacteria to resist viral attack. The term Restriction Enzyme originates from the studies of phage lambda. Restriction enzymes are used in biotechnology to cut DNA into smaller strands in order to study fragment length differences among individuals (Restriction Fragment Length Polymorphism – RFLP).