2. Nucleases are a class of enzymes called
hydrolases that catalyzes the hydrolysis
of nucleic acids(DNA,RNA) in all
organisms including plants and humans.
Nucleases are usually specific in action,
ribonucleases acting only upon
ribonucleic acids (RNA) and
deoxyribonucleases acting only upon
deoxyribonucleic acids (DNA).
There are two types of nucleases:
Endonucleases and exonucleases
3. Exonucleases degrade nucleic acids from
one end of the molecule. They opearate
either in 5’ 3’ or 3’ 5’ direction.
Endonucleases degrade nucleic acids at
specific internal sites, reducing it to smaller
and smaller fragments.
Restriction enzyme,a endonuclease due to
it’s cleavage at specific nucleotide
sequence find so much importance in
recombinant DNA technlology.
4. • Nuclease cleavage
sites
• (phosphodiester
linkage)
• Cleavage at bond A
generates a 5’-
phosphate and a 3’
OH terminus
• Cleavage at bond B
generates a 3’-
phosphate and a 5’-
hydroxyl terminus
5. restriction endonuclease
A restriction enzyme (or restriction
endonuclease) is an enzyme that cuts double-
stranded or single stranded DNA at specific
recogonition sequences.
These enzymes are found in bacteria.
Restriction enzymes are of four types.
Type 1and 2 require ATP to hydrolyze DNA.
Type 3 is independent of methylated
DNA,whereas type 4 targets methylated DNA..
6. Exonucleases degrade nucleic acids by
sequentially removing nucleotides from their
ends.
Two in common use are snake venom
phosphodiesterase and bovine spleen
phosphodiesterase.
Both enzymes act either on DNA or RNA.
But two enzymes have complementary
specifities.
7. Snake venom phosphodiesterase acts by ‘a’
cleavage and starts at the free 3’OH end of the
polypeptide chain,liberating nucleoside 5’
monophpsphates
8. In contrast to snake venom
phosphodiesterase,spleen enzyme acts by ‘b’
cleavage and starts at the 5’ end of a nucleic
acid releasing 3’nucleoside monophosphates.
9. Role of nucleases
Processes under control of nucleases are
protective mechanisms against "foreign"
(invading) DNA
degradation of host cell DNA after virus
infections
DNA repair
DNA recombination,
DNA synthesis
DNA packaging in chromosomes and viral
compartments, maturation of RNAs or RNA
splicing.
