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Difference Between Exonuclease and Endonuclease

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  • An endonuclease is a class of hydrolase that cleaves nucleic acids at the middle.
  • The action of endonucleases may result in two or more fragment of nucleic acids.
  • Endonucleases are capable of acting on both DNA and RNA.
  • The cleavage of some endonucleases such as deoxyribonucleases (DNases) is non-specific.
  • However, many endonucleases cleave the target nucleotide sequences in a specific manner. These type of specific endonucleases are called restriction endonucleases.


  • An exonuclease is a type of hydrolases that cleave the nucleic acid chain at its end.
  • Exonucleases remove nucleotides one by one from the nucleic acid chain by hydrolyzing the phosphodiester bonds at either 3’ or 5’ ends.
  • Three types of exonucleases can be identified in both prokaryotes and eukaryotes. They are 5’ to 3’ exonuclease, 3’ to 5’ exonuclease, and poly (A)-specific 3’ to 5’ exonucleases.

Difference Between Exonuclease and Endonuclease

DefinitionEndonuclease refers to an enzyme that cleaves the polynucleotide chain separating nucleotides other than the two end ones.Exonuclease refers to an enzyme that cleaves polynucleotide chain from the end of the chain by removing the nucleotides one by one.
Mechanism of ActionEndonucleases cleave nucleic acids at the middle of the nucleic acids.Exonucleases cleave nucleic acids at the ends.
Lag PhaseRestriction endonucleases undergo a lag period before their activity.Exonuclease does not have a lag period before their activity.
Blunt/Sticky EndsEndonucleases may form either blunt ends or sticky ends.Exonucleases form sticky ends.
ResultsEndonucleases result in oligonucleotides.Exonucleases result in single nucleotides or nucleosides.
RoleEndonucleases block the entry of pathogens.Exonucleases have no significant role in blocking the entry of pathogens.
SpecificitySpecific endonucleases, also called restriction endonucleases, are available that cleave specific sites within a DNA sequence.Exonuclease is usually non-specific.
Defensive propertiesEndonucleases have defensive properties against the entry of pathogenic microorganisms.Exonulceases do not have defensive properties.
Effect on circular DNARestriction endonuclease can cleave specific sites within a circular DNA.Exonulceases have less activity towards circular DNA as compared to linear DNA.
Free endsFree 3’ or 5’ ends are not necessary for the action of endonucleases.The ends should be free for the action of exonucleases.
InhibitionEndonucleases cannot be inhibited phosphorothioate bonds unless the entire sequence has the bonds between all nucleotides.Exonuclease can be inhibited by adding five phosphorothioate bonds in a row to a sequence.
ExamplesDNases, S1 nuclease, and restriction enzymes such as Bam H1, Hind III, and Eco RI are the examples of endonucleases.Snake venom, spleen phosphodiesterase, 3’ to 5’ exonuclease domain of the DNA polymerase III, the 5’ to 3’ exonuclease activity and the 3’ to 5’ exonuclease domain of the DNA polymerase I are examples of exonucleases.


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