Zinc fingers

Presented by
Vipin kumar
2014h129007p

Introduction
Discovered as repeating unit motif later
known as zinc finger as gripped and
grassped dna.
Tf3 of 5s rna gene,repeating structure:
a.30aa repeat in the sequence
b.Proteolysis results in limit digest of 3kda
structures
c.7-11 zn molecules attached

Tf3 a structure

Unique principle of dna

recognition
Instead of using 2 fold symmetry dna
helix,zn finger dna(or rna) binding module
linked tendemly in linear fashion.
Similar structural feature of ech unit but
achieve specific aafinity by varying key aa
residues.
This modular design allow to design large
no.of combination poosilities to recognise
sp.sequences.

AA residues involved in
interaction

Structure of zn finger
2 antiparallel sheet which contains loop
formed by 2 cys,and an helix containing
the
His-his loop
The two structural units are connected by
zn ion.
Dna recognition reside mostly in helical
region of str.
zn fingers are str.independent and linked
by flexible linker.

Modular recognition of dna

molecule
Zif268 TF dna binding domain having 3 zn
finger peptide model
1aa to 1bp linkages unlike other dna
binding motifs
Each finger interacts with 3 bp subsite,3 znf
motif recognizes 9bp sequence
Uses both statrands for recogniton
Highly specific in nature

Dna interaction

Zn finger affinity
Specificity of recognition of each zn finger
enable to improve specificity to higher level.
Main source of specificity -1,3,6 of
recognition helix,cross strand interaction at
2 influence specificity hence 4bp
overlapping subsites.
Selection of specific zn finger by phage
display method allow to design zn motif
with different specificity.

Improving znf affinity for

dna

9bp recognition znf motif linked to designed

18bp recognition zn motif via 2 ways 3*2 or 2*3
znf linkages.
Six fingers domain binding 18 bp target sites
18 bp sequences offers high specifity,one in
whole human genome (4^16=4.2*10^9)
3*2F picomolar affinity for dna and more
sensitive to mutation allow its preeference over
2*3f construct.
Offers single gene recognition tested on 20,000
dna sequence microarray

Some application of znf protien

Fusing poly zn finger motifs with variety of

effectors protiens (pabo and
collegues,sangamo Inc.)
Inhibition of HIV virus expression,zfp targeted
to hiv promotor terminal repeat,activated by
tat protien represses expression
Huntingtton desease elimination by
selectively differentiating mutant extended
repeats of CAG nts from wild type(no.of
repeats differ)

VEGF stimulation by ZNF

protiens

Activation of transcription factor VEGF by zfp invivo in

mouse
Vegf key regulator and effector in angiogenisis and
alternatively spliced.
Results in creation of new blood vessels by action on
promotor of VEGF gene
Splice variants ratio are naturally produced and no leakage
og vessels.

Zfp mediated gene correction

Homology directed recombination allow
gene editing mediated by znf.
Gene targetting by replacing defected gene
by hdr.
Chimeric zf make ds break and exogenous
right dna copy allow gene repalecement by
hdr.
Uses in gene therapy more efficiently by
targetting sp.dna sites to be replaced

Thank you