ZNF or Zinc Finger proteins A zinc finger is a small protein structural motif that is characterized by the coordination of one or more zinc ions (Zn2+) in order to stabilize the fold.

1. ZINC FINGER PROTEIN
By- Sanchit Dhankhar
1420923
MMCP

2.  Zinc finger motif- characteristics
 3D structural motif of the protein
 Characterized by the co-ordination of one or
more Zn ions
 Zinc ion is held in place by two cysteine and two
histidine R group
 Present in tandem repeats
 Interacting with DNA and RNA
 Zinc ion holding the structure together
 Each zinc motif consist of the 30 amino acid and
folds into beta-beta alpha structure
 Each finger primarily binds to a triplet within the
DNA substrate
2

3. History
 Zinc fingers were first identified in the Xenopus
Laevis in the laboratory of Aaron Klug.
 Revealed that the binding strength of the
transcription lllA is because of the presence of
the Zinc coordinating finger like structures.
3

4. Zinc finger motif
(functions)
These are the most abundant
proteins in the eukaryotes
genomes. There functions are
diverse and include;
 DNA recognition
 RNA packaging
 Transcriptional activation
 Regulation of apoptosis
 Protein folding and assembly
 Lipid binding
4

5. Zinc protein
families
 40 types of the zinc fingers in UniPortKB
Treble clef fingers Zinc Ribbons Cys2His2
5

6. Cys2His2
 Largest known DNA binding family in the multicellular
organisms
 Compound of 2 Beta layers and 1 Alpha helix
 Two cysteine and two histidine residues located in
certain positions bind zinc to stabilize the structure
 Plays role in – Development, differentiation and
suppression of malignant cell transformation
6

7. DNA Repair
Mechanism
 DNA repair mechanism
1. GENOME EDITING
2.ZINC FINGER NUCLEASE
3.COMPONENTS OF ZFN
7

8. ZFP’s
 Each ZFN consists of two functional domains-
a) A DNA binding domain comprised of a chain of two
finger modules.
b) A DNA-cleaving domain comprised of the nuclease
domain of Fok l
8

9. ZFP’s
 Designed to target any gene in any genome
 Delivered to the cell as DNA or RNA
 ZFN proteins are expressed
 Translocate to the nucleus
 Bind their target sites with high specificity
 Fok l nuclease form its catalytically active dimer
 Creates a single, specific double-strand break at the user
defined locus
 Living cells have evolved several methods to repair
double-strand breaks
 Endogenous processes can be harnessed to create gene
knockouts or knock-ins
9

10. Zinc finger nuclease- these are the
artificial restriction enzymes
 DNA binding domain-these are
composed of a tanden array of
multiple ZF modules in which each
ZF recognizes a 3 bp DNA subsite
 DNA cleavage domain- cleaving
domain of ZFNSs contains type ll
restriction enzyme Fokl
 ZFNs fuse the cleavage domain to
the C-terminuss of each zinc finger
domain
 Two cleavage domains must
dimerize and cleave DNA
10

11. Zinc finger nuclease
 Fok l type ll restriction enzyme-
Naturally found in Flavobacterium
okeanokoites, consisting of an N-
terminal DNA-binding domain and a
non-specific DNA cleavage domain
at the C-terminal
11

12. Applications of ZFN
 Functional genomic/Target validation
1. Creation of gene knockout in multiple cell lines
2. Complete knockout of gene not amenable by RNAi
 Cell-based screening
1. Creation of knock in cell line with promoters, fusion tags or reporters
integrated into ungenerous genes.
 Cell lines optimization
1. Creation of cell lines that produce high yield of proteins or antibodies.
12

13. THANKYOU
13