Question

Propose a chemical mechanism for the cleavage of double stranded DNA by a restriction enzyme. At a minimum, how many specific domain types must there be and what role does each play. Explain the role of any cofactors necessary in the reaction.

Answer 1

Restriction enzymes are a class of endonuclease group of enzymes. For cleavage of the DNA, all restriction enzymes make two incisions, once through each sugar-phosphate backbone of the DNA double helix in each strand. Restriction enzymes recognize a specific sequence of nucleotides and produce a double-stranded cut in the DNA. The recognition sequences can also be classified by the number of bases in its recognition site, usually between 4 and 8 bases, and the number of bases in the sequence will determine how often the site will appear by chance in any given genome, e.g., a 4-base pair sequence would theoretically occur once every 4*4 or 256bp, 6 bases, 4*6 or 4,096bp, and 8 bases would be 4*8 or 65,536bp. Many of them are palindromic, meaning the base sequence reads the same backwards and forwards.

All restriction enzymes require require AdoMet and ATP cofactors for their roles in DNA methylation and restriction digestion , respectively.