A PRESENTATION ON RESTRICTION ENDONUCLEASE (BgIII) Introduction Biochemical significance Structure of BglII Composition of active site Mechanism of action of BglII Conclusion Sources

1. A PRESENTATION ON RESTRICTION
ENDONUCLEASE (BgIII)
BY
GROUP 1
SULEIMAN ZAKARI (220000073)
NNAJI PRECIOUS CHINYERE
(220000082)
BCH 815

2. Outline
•Introduction
•Biochemical significance
•Structure of BglII
•Composition of active site
•Mechanism of action of BglII
•Conclusion
•Sources

3. Restriction enzyme also known as restriction
endonuclease, or restrictase are group of enzymes that
cleaves DNA into fragments at a specific restriction site
within its recognition sequence. To cut DNA, all
restriction enzymes make two incisions, once through
each sugar-phosphate backbone (i.e. each strand) of the
DNA double helix.
Introduction

4. Biochemical significance
Genetic Engineering: producing recombinant
DNA for variety of purposes including;
Vaccine development
Genetic recombination and recombinant DNA
repair

5. Structure
Name:
BglII Restriction
Endonuclease
EC Number: 3.1.21.4
Recognition sequence:
AGATCT
Fig 1: Crystal structure of BglII complexed with
DNA (Source; RCSB PDB)

6. Active Site
BglII's active site is similar to other endonucleases',
following the sequence Asp-(X)9-Glu-X-Gln.
In its active site there sits a divalent metal cation, most
likely Mg2+, that interacts with Asp-84, Val-94, a phosphoryl
oxygen, and three water molecules.

7. Mechanism of action of BglII
Fig 2: Steps involved in DNA cleavage by a BglII restriction endonuclease

8. BglII catalyses phosphodiester bond cleavage at the DNA backbone through
a phosphoryl transfer to water. This mechanism requires a base to generate
the hydroxide ion from water, which will act as the nucleophile and attack
the phosphorus in the phosphodiester bond. Also required a metal cation
Mg2+ to stabilize the extra negative charge of the pentacoordinated
transition state phosphorus by stabilizing the leaving group (3’-O−).
(Lukacs et al; 2000)

9. What type of mechanisms
involved?
Step 1: Proximity and orientation effect
Step 2: Covalent catalysis
Step 3: Electrostatic interactions or metal ion catalysis

10. Rethink
A question could come into your mind like “If a bacteria
produces this kind of enzyme then how can they protect
their own DNA from being cut by the restriction enzymes?
RESTRICTION MODIFICATION SYSTEM

11. Conclusion
Restriction endonucleases play a very important role in
modern molecular cloning techniques. Because of their
unique recognition/cut sites, restriction enzymes can be
used to precisely cut DNA at specific locations in a
predictable manner.

12. References
•Lukacs CM, Kucera R, Schildkraut I, Aggarwal AK (February
2000). "Understanding the immutability of restriction enzymes:
crystal structure of BglII and its DNA substrate at 1.5 A
resolution". Nature Structural Biology. 7 (2): 134–40.
doi:10.1038/72405. PMID 10655616. S2CID 20478739
•RCSB Protein Data Bank 2022. Retrieved at
https://www.rcsb.org/structure/1DFM. Accessed on 30th
October, 2022.