Hello friends, in today's article we see Restriction Endonucleases Enzymes Information, and their different types with examples.
so let's start
Restriction Endonucleases:
so first arise is What is restriction endonucleases?
Ans: Restriction endonucleases are the type of Enzymes, but it has specificity in restriction for cleavage.
Discovery of Restriction Endonucleases:
The Restriction endonucleases were first found from the studies of Phage Lambda.
In 1960, Werner Arber and Mathew Meselson studies the type I restriction Enzymes.
In 1970, Hamilton O. Smith, Thomas Kelly, and Kent Wilcox isolated the first type II restriction enzymes and then characterized the restriction enzymes type II.
This enzyme is isolated from the Bacterium Haemophilus Influenzae.
For their work in the discovery and Characterization of Restriction enzymes, in the 1978, they got the noble prize for Physiology or Medicine was awarded to Werner Arber, Daniel Nathans, and Hamilton O. Smith.
Classification of Restriction Endonucleases:
Restriction endonucleases are enzymes that produce internal cuts, called Cleavage, in the DNA/RNA Molecules
Restriction Endonucleases (Restriction Enzyme) is a bacterial enzyme that cuts dsDNA into fragments.
This process has done after recognizing a specific Nucleotide sequence known as Recognition or Restriction site.
Restriction Enzymes are beloved to be evolved by bacterial to resist Viral attacks.
Restriction Enzymes are also known as Molecular Scissor.
They are Classified Based Upon:
1) Their Composition
2) Their Enzyme Cofactor requirement
3) The Nature of their Target Sequence
4) Position of their DNA Cleavage site
5) Relative to the target Sequence
Nomenclature of Restriction Endonucleases:
At present time, we have discovered (identified) more than 3000 restriction endonucleases.
Each and Every Enzyme give a specific and definite name.
Enzymes nomenclature is based on the bacterium from which it was isolated.
The First Three Letters of the enzyme's name are derived from the First Letter of the Genus name and the First Two Letters of Species Name.
Since each bacterium may contain several different restriction enzymes a Roman Numeral is also used to Identify each Enzyme.
In Case, Enzyme is isolated from different strains of the same bacterial first Letter of the strain is used to identify each enzyme.
For example, EcoRI is derived from E for Escherichia ( Genus), Coli (species) and R for Strain, and lastly I ( first identified from bacterium).
A few more derivations of the same Restriction enzyme names are given in the following table below
How They Work:-
Restriction Enzymes recognize a specific site of Nucleotide Sequence.
And They produce a double-stranded cut in the DNA.
These cuts are divided into two types
1) Blunt Ends
2) Sticky ends
1) Blunt Ends:-
- Blunt Ends cut is when A Straight cut, down through the DNA that results in a Flat pair of Bases on the Ends of the DNA.
- The Blunt Ended Fragments can be joined to any other DNA fragment with Blunt Ends only.
- Enzymes Useful for certain types of DNA Cloning Experiment
2) Sticky Ends:
- Staggered end is on DNA molecules with short, single-stranded Overhangs.
- the DNA Fragments with Complimentary sticky ends can be combined to create new molecules which allow the creation and manipulation of DNA Sequences from different sources.
Types of Restriction Endonucleases:
At present day, More than 2500 different restriction endonuclease have been identified that are isolated from a number of species of Bacteria.
More than 3000 Restriction Enzyme are now available for use in the Lab.
These enzymes belong to three different classes of Restriction Endonucleases that can be distinguished from each other on the basis of their structure, cofactors required, and features of their restriction and cleavage site.
Restriction enzymes are categorized into three general groups.
1) Type I Restriction Endonucleases
2) Type II Restriction Endonucleases
3) Type III Restriction Endonucleases
1) Type I Restriction Endonucleases:
- Type I restriction Enzyme have a complex structure with three different subunits
a. Endonucleases
b. Methyl Transferase
c. Recognition
- These enzymes cut DNA molecules at a Random site, far, (= 1kb) from the recognition sequence.
- Their restriction sites are also complex and discontinuous with Spacers.
- They may have biological significance but are of very little practical value since the site where the DNA is going to be digested can not be predicted.
- Type I Restriction Endonucleases enzymes are complex endonucleases and have a recognition sequence of 15bp.
- They cleave the DNA molecule about 1000 bp away from the strand of the Sequence "TCA"(TCA is not a cycle of glycolysis is a sequence of DNA) located within the recognition site,
- Eg. EcoK-12, EcoB, etc
- The type I Restriction endonucleases enzyme work with the help of many co factor-like as S-adenosyl Methionine, Hydrolysed Adenosine Triphosphate ( ATP), and Magnesium (Mg 2+)
- The Type I Restriction Enzyme methylases, The DNA Molecule sequence at the site of recognition.
- The type I Restriction enzyme translocate the DNA Sequence.
Subunits of Type I Restriction endonucleases:
Type I restriction enzyme possesses three subunits::
1) HsdR:- is required for Restriction
2) HsdM:- is necessary for adding methyl groups to host DNA ( Methyl Transferase Activity)
3) HsdS:- is Important for the specificity of cut site recognition in addition to its methyltransferase activity.
2) Type II Restriction Endonucleases:
- Types- II restriction endonucleases enzyme cleave DNA molecules at very precise and defined positions close to or within the recognition sequence.
- These enzymes require magnesium (Mg 2+) ion as a cofactor for their activity.
- They are smaller in size, in comparison to type-I and type-III restriction endonucleases enzymes.
- They are mostly bound to DNA molecules as Homodimers.
- The recognition site is mostly symmetric and shows two-fold symmetry.
- Most of the Type II enzymes is cleave within the symmetric recognition sequence, Eg. HhaI, HindII, and Not I, but
- There are other classes also that cleave outside their recognition sequence.
- A Few another type II restriction enzymes clean on both sides of the recognition sequence releasing a small fragment.
- The Type II restriction enzyme are remarkably stable and induce cleavage either, in most cases, within or outside their recognition sequences, which are symmetrical.
- The Type II restriction Enzyme work with the help of one a single cofactor like Magnesium (Mg 2++)
- The 1st Type II Enzyme to be isolated was HindII in 1970.
- Only Type II restriction endonucleases are used for restriction mapping and gene cloning also, in view of their cleavage only at specific sites.
- Type II restriction enzymes can generate two different types of cuts depending on Whatever they cut both strands at the center of the recognition sequence.
- The former cut will generate " Blunt ends " with no nucleotide overhangs
- the latter, generate " Sticky" or " Cohesive Ends "
Subunits of Type II Restriction Endonucleases:
these subgroups are defined using letter suffixes.
1) Type II B Restriction Endonucleases
2) Type II E Restriction endonucleases
3) Type II M Restriction endonucleases
4) Type II T Restriction endonucleases
3) Type III Restriction endonucleases:-
- Type III restriction enzymes are a group of endonucleases that recognize a non-palindromic sequence, comprising two inversely oriented sites.
- Type II Restriction Endonuclease is infrequently used in molecular biology, as they have few relevant applications.
- They are less well characterized than Type II restriction enzymes
- The type III Restriction enzymes themselves consist of two proteins that function as a single protein complex: R protein, and Restriction enzymes.
- Type III enzymes recognize a 5-6 base, Non-palindromic sequence, and require 2 inversely oriented recognition sites for cleavage.
- The Type III restriction enzyme slides along the DNA Molecules and upon encountering a second complex cleaves downstream of the recognition site, typically 25-28 bases away.
- Type III restriction enzymes require ATP, S-adenosyl methionine, and Magnesium ( Mg 2++) ions.
- Eg. EcoP151 is currently the only type III restriction enzyme commercially available.
this is all about the Restriction of endonucleases.
If you want to know about the Father of Indian Biotechnology, then buy this book, from the following link
Click:- To Buy
Read More Articles
No comments:
Post a Comment
problem related comment