I was wondering how a restriction enzyme creates sticky end (or blunt end)? What makes it cut so? Thanks in advance.
According to Pingoud and Jeltsch (2001):
Enzymes that produce blunt ends or sticky ends with 3′-overhangs approach the DNA from the minor groove side and mainly use a β-strand and β-like turn for DNA recognition.
Enzyme that produce sticky ends with 5′-overhangs contact the DNA from the major groove side and mainly use an α-helix and a loop for DNA recognition.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC55916/
Restriction enzymes refers to a group of endonucleases which cleaves the DNA at specific points known as recognition sequences or sites. The mechanism of restriction enzymes lies in the property that it cleaves the DNA sequence within their specific recognition sites.
Recognition sites are specific area or sequences in the genetic molecule which these enzymes recognize as sites for cleavage. Recognition sites are unique for different restriction endonucleases. For type II restriction endonucleases, recognition sites are mostly palindromic sequences with rotational symmetry. DNA has a double stranded helical structure where, the nucleotides of the two strands of DNA are complementary to each other. There are certain sequences in such a structure where, the first half of the sequence is a mirror image of the second half of the complementary strand and reads identical from same end. Such sequences are termed as palindromic sequence with rotational symmetry.
Eg:-
5’GAATTC3’
3’CTTAAG5’
The restriction endonuclease moves along the surface of the DNA until it recognises its target sites. After recognition, it initiates DNA binding in the presence of Mg2+ ions resulting in cleavage at specific sites.
Length of the recognition sequence dictates how frequently the enzyme will cut in a random sequence of DNA. Enzymes with a 6 bp recognition site will cut, on average, every 46 or 4096 bp; a 4 bp recognition site will occur roughly every 256 bp.
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