InterPro: IPR011578 Restriction endonuclease, FokI, C-terminal/endonuclease I, core

There are four classes of restriction endonucleases: types I, II,III and IV. All types of enzymes recognise specific short DNA sequences and carry out the endonucleolytic cleavage of DNA to give specific double-stranded fragments with terminal 5'-phosphates. They differ in their recognition sequence, subunit composition, cleavage position, and cofactor requirements [1, 2], as summarised below:

• Type I enzymes (EC:3.1.21.3) cleave at sites remote from recognition site; require both ATP and S-adenosyl-L-methionine to function; multifunctional protein with both restriction and methylase (EC:2.1.1.72) activities.
• Type II enzymes (EC:3.1.21.4) cleave within or at short specific distances from recognition site; most require magnesium; single function (restriction) enzymes independent of methylase.
• Type III enzymes (EC:3.1.21.5) cleave at sites a short distance from recognition site; require ATP (but doesn't hydrolyse it); S-adenosyl-L-methionine stimulates reaction but is not required; exists as part of a complex with a modification methylase methylase (EC:2.1.1.72).
• Type IV enzymes target methylated DNA.

Type II restriction endonucleases (EC:3.1.21.4) are components of prokaryotic DNA restriction-modification mechanisms that protect the organism against invading foreign DNA. These site-specific deoxyribonucleases catalyse the endonucleolytic cleavage of DNA to give specific double-stranded fragments with terminal 5'-phosphates. Of the 3000 restriction endonucleases that have been characterised, most are homodimeric or tetrameric enzymes that cleave target DNA at sequence-specific sites close to the recognition site. For homodimeric enzymes, the recognition site is usually a palindromic sequence 4-8 bp in length. Most enzymes require magnesium ions as a cofactor for catalysis. Although they can vary in their mode of recognition, many restriction endonucleases share a similar structural core comprising four beta-strands and one alpha-helix, as well as a similar mechanism of cleavage, suggesting a common ancestral origin [3]. However, there is still considerable diversity amongst restriction endonucleases [4, 5]. The target site recognition process triggers large conformational changes of the enzyme and the target DNA, leading to the activation of the catalytic centres. Like other DNA binding proteins, restriction enzymes are capable of non-specific DNA binding as well, which is the prerequisite for efficient target site location by facilitated diffusion. Non-specific binding usually does not involve interactions with the bases but only with the DNA backbone [6].

This entry represents FokI restriction endonucleases, as well as bacteriophage endonucleases I (Holliday junction resolvase). FokI recognises the double-stranded DNA sequence 5'-GGATG-3' and cleaves DNA phosphodiester groups 9 base pairs away on this strand and 13 base pairs away on the complementary strand [7]. Endonuclease I (EC:3.1.21.2) is a junction-resolving enzyme encoded by Bacteriophage T7, that selectively binds and cleaves four-way Holliday DNA junctions [7]. The structure of the enzyme shows that it forms a symmetric homodimer arranged in two well-separated domains. Each domain, however, is composed of elements from both subunits, and amino acid side chains from both protomers contribute to the active site [8].