Restriction Enzymes

Introduction

Restriction enzymes are also called restriction endonucleases or re+9strictase. These enzymes are specific proteins that are produced by bacteria or other cellular organisms that can help to cleave or break up DNA from a particular sequence. These restriction enzymes are very specific to the site where they act; in fact, there are major roles of these enzymes in DNA cleaving. These enzymes can also be helpful in cleaving foreign DNA particles which can help in specific recombinant DNA technology.

History of Restriction Enzymes

Restriction enzymes were originally obtained from bacteriophages. Bacteriophages are certain bacteria infecting viruses. The restriction enzymes were discovered by Werner Arber. These enzymes were first noticed in the laboratory of Salvador Luria, Jean Weigle, and Giuseppe Bertani. The discovery took place in 1950, and during that time, the bacteriophage grew on a single strain of Escherichia coli. 

Different types of restriction enzymes were obtained or discovered at different years. The type 1 restriction enzyme, which actually cleaves DNA randomly away from its restriction site, was found in the 1970s. Type 1 Restriction enzymes were discovered by Arber and Meselson. Type 1 restriction enzymes are also known as endonuclease enzymes. 

The type 2 restriction enzyme was found by Hamilton O. Smith, Thomas Kelly, and Kent Wilcox. Note that recombinant DNA technology is only possible due to restriction enzymes. 

These enzymes first came into existence when the main restriction enzyme was extracted and further was used on the DNA of bacterias in order to produce special genetic sequences. Many new drugs are being manufactured due to the enzyme itself. 

Important Term Regarding Restriction Enzymes

Recognition site- One of the characteristics of recognition enzymes is the fact that they can only act on specific areas of a DNA sequence. Different recognition sites have been reported by the discoverers and researchers. These sites have both pharmaceutical and research applications.

Palindromic sequence- It refers to a nucleic acid sequence in RNA or DNA molecules that are double-stranded. In this, a particular direction’s reading on any one of the strands is always the same as the complementary strand’s sequence having the same direction. 

Types of Restriction Enzymes

As mentioned above, there are numerous restriction enzymes discovered to date. There are different enzymes for humans and different ones for other species. Since the main function of restriction enzymes in biotechnology is to work with recombinant DNA, these enzymes are classified according to their restriction sites and even according to the mode of their action. 

Many restriction enzymes like to cut at the open end, while others like to operate on the closed ones. You can also see restriction enzymes working on special repeating sequences. All these will open up the DNA in a special way that becomes very specific for operation. 

Restriction enzymes are also known on the basis of organisms from which they are extracted or from which they are generated. The special functions of these enzymes are also dependent on the bacteria from which they are removed.

Naturally occurring restriction enzymes are classified into four groups 

1. Type I enzymes- These restriction enzymes are basically responsible for cutting DNA at any remote site from a recognition site. These restriction enzymes are activated by the special ATP and S-adenosyl-L-methionine. These enzymes are pretty special and have multifunctional activities such as restriction digestion and methylase activities.
2. Type II enzymes- This type of restriction enzyme can cleave or break the DNA from a site that is near the actual recognition site. It does require a high amount of magnesium for its function. It is operative for single-use, and this enzyme can work without the presence of methylase.
3. Type III enzymes- This restriction enzyme also breaks the DNA at a site near the actual recognition site. It requires ATP for the working, but it does not require any hydrolase. S-adenosyl-L-methionine is needed in the reaction to start the reaction. But once the reaction proceeds, it has nothing to do with the working of enzymes. This restriction enzyme can help in the digestion of the DNA with a modification methylase.
4. Type IV- The type IV restriction enzyme is a special endonuclease enzyme that only acts on modified DNA. This restriction endonuclease acts on the DNA and is widely used in the field of biotechnology. Examples of restriction enzymes are methylated and hydroxymethylated. 
5. Type V- The Type V restriction enzyme is not basically a reaction enzyme for DNA. This restriction endonuclease acts on RNA and is initiated with an RNA guide known as the gRNAs.

Artificial Restriction Enzymes

Artificial restriction enzymes are special restriction enzymes that are not found in biological systems. These restriction enzymes are a product of biotechnology and are made up of bioengineering. Such artificial restriction enzymes can be made by using or joining a natural DNA restriction enzyme along with the special engineer restriction enzyme, which is a product of biotechnology. 

These restriction enzymes are made by special biotech methods, and on the other hand, they can target huge DNA sites with more than 36 base pairs. In the year 2013, a new technology known as CRISPR-Cas9 was invented, which is a more powerful engineer for editing the genome. 

Special artificial ribonucleases are also available these days, which can act as restriction enzymes for the RNA. These enzymes also have the selective potential to cleave the DNA on one side, and it is sure that you don’t get a mismatch for two possible cleavage sites.

Nomenclature of Restriction Enzymes

Restriction enzymes were discovered earlier in 1970, and people didn’t know how to differentiate among them. From the time of their discovery, restriction endonuclease has been named according to different findings, but one needs to have proper knowledge of nomenclature so that it doesn’t create confusion. There are more than 3500 types of type 2 restriction enzymes characterized these days. Basically, all these enzymes are named according to the bacteria from which it was isolated. The naming system is based on the genus and species of the strains. For example, EcoR1 is named accordingly as mentioned below – 

• E- the E here denotes the genus name, which is basically the Escherichia

• Co- This denotes the specific species of Escherichia. Here, the co marks coli

• R- It denotes the strain of the bacteria. The RY13 strain is denoted here

• 1- This one means that this enzyme is identified first among all endonucleases of the species

Application of Restriction Endonucleases or Restriction Enzyme

Restriction enzymes are isolated from different bacterias and living organisms. These enzymes are used to manipulate DNA and have thousands of applications. 

• The main role of these enzymes is basically to assist the input or the insertion of a specific sequence of a gene or a DNA into the circular chromosomal material known as a plasmid. The plasmid has been used in recombinant DNA technology with gene cloning

• Different restriction enzymes are also efficient in digesting genomic DNA from genes. DNA analysis could be used for the southern blots. Such blotting techniques are used in delivering the specific DNA patterns, which could further be used in DNA fingerprinting

• The creation of vaccines has also been possible due to restriction enzymes. There is literally no chance of DNA alteration and production of vaccines without the use of restriction enzymes

Examples of Restriction Enzymes

Some well-known examples of restriction enzymes include HhaI, HindIII, and NotI. These kinds of enzymes come under the principal restriction enzymes available commercially.

Conclusion

Above in the article, we have mentioned all the necessary information one needs to learn about restriction enzymes. Restriction enzymes or endonucleases are among the most used ones in the world of biotechnology. The different types of restriction enzymes can cleave DNA in specific manners, and this could be useful in using it with plasmids, which are the particular chromosomal matter of bacterias.