What is a RESTRICTION ENZYME?

A restriction enzyme is a protein that recognizes
a specific, short nucleotide sequence and cuts the
DNA only at that specific site, which is known
as restriction site or target sequence.
What is the function of a restriction endonuclease?
In cells restriction endonucleases are used to
cut up invading DNA from viruses.
The restriction sites are ones found in viral DNA.
If, when a virus injects DNA in a bacterium,
the restriction enzymes cut up the viral DNA then
it cannot take over the bacterium and cause it to
manufacture more viruses.

How do restriction enzymes cleave DNA?
A major protective strategy for the host is to
use restriction endonucleases(restriction
enzymes) to degrade the viral DNA on its
introduction into a cell. These enzymes recognize
particular base sequences, called recognition
sequences or recognition sites, in their
target DNA and cleave that DNA at defined
positions.

and if both adenines are un-methylated (a signal that the DNA is non-host DNA). which makes them important molecular motors. and can be hundreds of bases away. compare the methylation status of two adenines within the recognition sequence.Types of Restriction Enzymes: Restriction enzymes are classified biochemically into three types. Type I and III systems. Type II. Although these enzymes recognize specific DNA sequences. but also due to other factors). Both require ATP for their proper function. both the methylase and restriction activities are carried out by a single large enzyme complex. A major type of Type II enzymes are sometimes referred to as Type IV enzymes. the sites of actual cleavage are at variable distances from these recognition sites. and Type III. These are designated as Type I. The cleavage of DNA appears to occur after blockage of the translocation activity (often following collision with another translocating Type R-M enzyme. These enzymes read the methylation status of their recognition sequence. Type I restriction enzymes produce DNA cleavage following translocation of the DNA. the enzyme undergoes a conformational switch that .

HindIII and NotI that cleave DNA within their recognition sequences. and type lIb enzymes cut sequences twice at both sites outside the recognition sequence.turns the enzyme into a molecular motor and endonuclease. while type IIa enzymes recognise non-palindromic sequences and cleave outside of the recognition site. The vast majority of known restriction enzymes are of type II. The most common type II enzymes are those like HhaI. and are useful for DNA analysis and gene cloning. The first to be discovered and utilized was EcoRI. However. Type II enzymes are further classified according to their recognition site. . Type lIs enzymes cleave the DNA at a considerable offset from the recognition sequence. and it is these that find the most use as laboratory tools. Most type II enzymes cut palindromic DNA sequences. which is staggered and its recognition sequence is 5′GAATTC-3′. the restriction enzyme is independent of its methylase. They produce discrete bands during gel electrophoresis. if either one of the adenines is methylated (a signal that the DNA is host DNA) then the enzyme acts as a maintenance methylase and methylates the other adenine. and cleavage occurs at very specific sites that are within or close to the recognition sequence. In Type II systems.

In other systems. of a larger. restriction-and-modification enzyme.Restriction enzymes usually occur in combination with one or two modification enzymes (DNAmethyltransferases) that protect the cell’s own DNA from cleavage by the restriction enzyme. they methylate one of the bases in each of the DNA strands. or as separate domains. a restriction enzyme and its “cognate” modification enzyme(s) form a restrictionmodification (R-M) system. Because there are only so many ways to arrange the four nucleotides—A. The methyl groups protrude into the major groove of DNA at the binding site and prevent the restriction enzyme from acting upon it. combined. Modification enzymes recognize the same DNA sequence as the restriction enzyme that they accompany. Together. In some R-M systems the restriction enzyme and the modification enzyme(s) are separate proteins that act independently of each other.C. the two activities occur as separate subunits. Restriction Enzymes as Tools: Recognition sequences typically are only four to twelve nucleotides long. but instead of cleaving the sequence.G .

As a result. recognition sequences tend to “crop up” by chance in any long sequence.and T–into a four or eight or twelve nucleotide sequence. Furthermore. and then gel electrophoresis is performed on this digest. So no matter the context in which a gene naturally appears. there is probably a pair of restriction enzymes that can cut it out. the sequences of some artificial plasmids include a “linker” that contains dozens of restriction enzyme recognition sequences within a very short segment of DNA. this restriction enzyme can be used to genotype the sample without completely sequencing it. potential “restriction sites” appear in almost any gene owr chromosome. and which will produce ends that enable the gene to be spliced into a “plasmid”. restriction enzymes specific to hundreds of distinct sequences have been identified and synthesized for sale to laboratories. the alternate nucleotide of the SNP causes the restriction site to no longer exist within the section of DNA). . Meanwhile. Another use of restriction enzymes can be to find specific SNPs. If a restriction enzyme can be found such that it cuts only one possible allele of a section of DNA (that is. The sample is first run in a restriction digest to cut the DNA.

They recognize short. If the sample is homozygous for the rarer allele. If the sample is heterozygous at that SNP. they cut DNA into fragments of a size suitable for cloning. Restriction enzymes have two properties useful in recombinant DNA technology. there will be three bands of DNA. First. This is an example of restriction mapping. many restriction enzymes make staggered cuts that create single-stranded sticky ends conducive to the formation of recombinant DNA. in . usually palindromic. because the cut will have occurred at the restriction site. the result will be two bands of DNA.If the sample is homozygous for the common allele. because it will not have been cut. How is restriction enzymes used in recombinant DNA technology? MESSAGE. sequences of 4–8 bp and. the sample will show only one band. Second. How many restriction enzymes are there? More than 3000 type II restriction endonucleases have been discovered.

The fourth letter typically comes from the bacterial strain designation. Restrictions sites in the viral genome (a "happy accident" of nature. the Roman numeral indicates the order in . even if it has the target sequences ordinarily targeted by them. This is because the bacterial restriction sites are highly methylated. as far as the bacteria are concerned. making them unrecognizable to the restriction enzyme. For example. A bacterium is immune to its own restriction enzymes. strain Rd. Restriction enzymes are named based on the organism in which they were discovered. the enzyme Hind III was isolated from Haemophilus influenzae. The orthodox type II enzymes are homodimers which recognize palindromic sites. fragmenting and destroying the DNA of invading bacteriophages before it can incorporate into the host's genome and take over the cell.the presence of Mg2+. since they don't appear to have any specific function in the virus) are cleaved by the bacterium's restriction enzymes. The first three letters of the name are italicized because they abbreviate the genus and species names of the organism. More than 400 restriction enzymes have been isolated from the bacteria that manufacture them. restriction enzymes function to defend the cell against invading viral bacteriophages. In live bacteria. Typically. The Roman numerals are used to identify specific enzymes from bacteria that contain multiple restriction enzymes. cleave the DNA within or in close proximity to the recognition sequence.

which restriction enzymes were discovered in a particular strain.Dna. (ii) 3′ overhangs: Again. . we see asymmetrical cutting within the recognition site. Type.Nomenclature And Classification . Kpnl cuts in this manner. BamHI cuts in this manner.jrank. Read more: Restriction Enzymes .html#ixzz4UhSZewQI Patterns of DNA Cutting by Restriction Enzymes: (i) 5′ overhangs: ADVERTISEMENTS: The enzyme cuts asymmetrically within the recognition site such that a short single-stranded segment extends from the 5′ ends. and Sequences .JRank Articles http://medicine. but the result is a single-stranded overhang from the two 3′ ends.org/pages/2778/Restriction-EnzymesNomenclature-Classification. Recognition.

Smal is an example of an enzyme that generates blunt ends. . The 5′ or 3′ overhangs generated by enzymes that cut asymmetrically are called sticky ends or cohesive ends. because they will readily stick or anneal with their partner by base pairing.(iii) Blunts: Enzymes that cut at precisely opposite sites in the two strands of DNA generate blunt ends without overhangs.

They proposed that the enzyme names should begin with a three-letter acronym in which the first letter was the first letter of the genus from which the enzyme was isolated and the next two letters were the first two letters of the species name. but as more enzymes have been found. . Extra letters or numbers could be added to indicate individual strains or serotypes. we now know that each major type of enzyme can contain subtypes. the enzyme Hindll was one of four enzymes isolated from H aemophilus in fluenzae serotype d. In addition. these schemes were very useful. and III (1. a formal proposition for naming the genes encoding REases and MTases was adopted (4). In this paper we revisit the naming conventions and outline an updated scheme that incorporates current knowledge about the complexities of these enzymes. When there were only a handful of enzymes known. We describe a set of naming conventions for REases and their associated MTases. II.The first three letters of the name were italicized.here are three main groups of restriction endonucleases (REases) called Types I. REases and DNA methyltransferases (MTases) have been named based on an original suggestion by Smith and Nathans (3). Later. considerable laxity in naming conventions has appeared. Thus. of which more than 3500 have been characterized (5). often from different genera and species with names whose three-letter acronyms would be identical. This especially applies to the Type II enzymes.2). Since 1973.