How Do Nucleases Help in DNA and RNA Processing?
Nuclease is the term used for an enzyme that cleaves nucleic acids. They are also known as nucleodepolymerase or polynucleotide. Nucleases belong to the enzyme class hydrolases and perform a specific role. The ribonucleases work on the ribonucleic acids (RNA) and deoxyribonucleases work on the deoxyribonucleic acids (DNA).
Some enzymes that have a general role to play such as phosphodiesterases, which hydrolyze phosphoric acid esters, can also be termed nucleases because nucleic acids are affected by their action. Nucleases are present in plants as well as animals.
Function of Nucleases
Nuclease has the role of cleaving the phosphodiester bonds between nucleotides of nucleic acids and causing single and double-stranded breaks in their target molecules. They are essential in living organisms for their several DNA repair aspects. If there are defects in certain nucleases, it can cause immunodeficiency or genetic instability. Nucleases also play a critical role in molecular cloning.
Types of Nucleases
Nuclease can be classified into two broad categories based on the locus of activity: Exonucleases and Endonucleases. While Exonucleases digest nucleic acids from the ends, the Endonucleases work on regions in the centre of target molecules. These categories can be further subcategorized into deoxyribonucleases and ribonucleases. The deoxyribonucleases (DNA nuclease) act on DNA, while the ribonucleases work on RNA.
Micrococcal Nuclease
Micrococcal nuclease is a type of endonuclease that primarily digests single-stranded DNA or RNA, specifically at AU- or AT-rich regions. This enzyme also digests double-stranded DNA or RNA and is a critical component of chromatin immunoprecipitation (ChIP) assays. Micrococcal Nuclease can digest 5'-phosphodiester bonds of RNA and DNA, and yields 3'-phosphate mononucleotides and oligonucleotides. It needs Ca2+ as a cofactor for performing its function and gets completely inactivated by EGTA or EDTA.
Mung Bean Nuclease
Mung bean nuclease is a single-stranded (ssDNA or RNA) endonuclease that helps in the removal of single-stranded extension in double-stranded DNA. It can be used for removing both 3' and 5' single-stranded overhangs from double-stranded DNA for the creation of blunt ends. It cleaves single-stranded RNA and DNA, cleaves the single-stranded region in a DNA hairpin and helps in the mapping of RNA transcripts.
Site Recognition
Before it can cleave the molecule, a nuclease must associate with a nucleic acid. It requires a certain degree of recognition and nucleases use both specific and nonspecific associations in their means of binding and recognition. Both modes or means play a critical role in living organisms, especially in the area of DNA repair.
Nonspecific endonucleases that are involved in DNA can scan DNA for damage or target sequence. This type of nuclease diffuses along with the DNA until it meets a target. Then, the residues of its active site mingle with the DNA chemical groups. For endonucleases such as BamHI, EcoRV, and PvuII, electrostatic interactions between the DNA and minimal surface area of the protein are involved in the nonspecific binding. The overall shape of the DNA is undeformed because of this weak association, thus remaining in B-form.
Site-specific nucleases can form associations that are much stronger in contrast. They can draw DNA into the deep groove of their DNA-binding domain. It causes a significant deformation of the DNA tertiary structure and is accomplished with basic residue-rich or positively charged surfaces. Such nucleases engage in electrostatic interaction with the DNA extensively.
Some nucleases involved in DNA repair are partial sequence-specific. The majority of the nucleases however are nonspecific and recognize structural abnormalities generated in the DNA backbone with the use of base pair mismatches.
FAQs on What Is a Nuclease? Definition, Types & Functions
1. What is a nuclease and what is its primary function in biology?
A nuclease is an enzyme that plays a crucial role in breaking down nucleic acids, such as DNA and RNA. Its primary function is to cleave the phosphodiester bonds that link nucleotide subunits together, effectively cutting the nucleic acid chain into smaller fragments. This action is fundamental to many cellular processes.
2. What are the main types of nucleases and how do they differ in their action?
Nucleases are broadly classified into two main types based on how they act on a nucleic acid strand:
- Exonucleases: These enzymes remove nucleotides one by one from the ends (either the 5' or 3' end) of a DNA or RNA molecule. They work like trimming the edges of a rope.
- Endonucleases: These enzymes cut the nucleic acid strand at internal sites within the molecule, rather than from the ends. They act like cutting a rope in the middle. Restriction enzymes are a well-known example of endonucleases.
3. What are restriction enzymes and why are they often called 'molecular scissors'?
Restriction enzymes, or restriction endonucleases, are a special class of nucleases that recognise and cut DNA at specific nucleotide sequences known as restriction sites. They are called 'molecular scissors' because of their precise ability to cut DNA at these defined locations, a fundamental technique used in recombinant DNA technology and genetic engineering to cut and paste pieces of DNA.
4. How is the action of a nuclease different from that of a DNA helicase?
While both enzymes interact with DNA, their functions are fundamentally different. A nuclease breaks the phosphodiester bonds of the DNA backbone, causing a cut in the strand itself. In contrast, a DNA helicase breaks the hydrogen bonds between complementary base pairs to unwind and separate the two DNA strands, without breaking the backbone. Think of helicase as unzipping a zipper, while a nuclease is like a pair of scissors that cuts the zipper tape.
5. What role do nucleases play in the human digestive system?
In the digestive system, the pancreas secretes nucleases into the small intestine to break down the nucleic acids (DNA and RNA) present in the food we eat. Specifically, deoxyribonuclease (DNase) digests DNA, and ribonuclease (RNase) digests RNA. This process breaks them down into smaller nucleotides, which can then be absorbed by the body and used for its own cellular needs.
6. Beyond genetic engineering, what is the importance of nucleases in a living cell's daily functions?
Nucleases are vital for maintaining cellular health and integrity in several ways:
- DNA Repair: They are essential for identifying and removing damaged or incorrect sections of DNA. Exonucleases can 'proofread' newly synthesized DNA and remove mismatched bases.
- Cellular Defense: Many bacteria use restriction nucleases to recognise and destroy the DNA of invading viruses (bacteriophages), acting as a primitive immune system.
- Apoptosis: During programmed cell death (apoptosis), nucleases are activated to systematically break down the cell's DNA.
7. Why do bacteria produce so many different types of restriction nucleases like EcoRI and HindII?
Bacteria produce a vast array of restriction nucleases as a defence mechanism against viruses. Each specific enzyme (like EcoRI from Escherichia coli or HindII from Haemophilus influenzae) recognises a unique DNA sequence. This diversity provides a broad defence, as different viruses will have different DNA sequences. The bacterium protects its own DNA from being cut by modifying it at these same recognition sites, a process called methylation, making its own DNA invisible to its nuclease enzymes.
8. What is an example of a nuclease that can degrade both DNA and RNA?
Benzonase is an example of a genetically engineered, non-specific endonuclease that can degrade all forms of both DNA and RNA (single-stranded, double-stranded, linear, and circular). Because of its broad activity and high efficiency, it is widely used in laboratory and industrial settings to remove nucleic acid contamination from protein preparations, which reduces viscosity and improves the purity of the final product.
