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DNA: Structure, Dynamics and
Recognition
L4: DNA deformation
Les Houches 2004
BASE PAIR OPENING
Bond vibrations
Sugar repuckering
DNA bending
1 fs
1 ps
1 ns
(10-15 s)
(10-12 s)
(10-9 s)
Domain movement
Base pair opening
1 s
1 ms
(10-6 s)
(10-3 s)
Transcription
Protein synthesis
Protein folding
RNA lifetime
2.5 ms / nucleotide
6.5 ms / amino acid
~ 10 s
~ 300 s
Biological time scale
Enzymatic base chemistry
HN3 imino proton
S
Adenine-Thymine base pair
S
HN1 imino proton
S
S
Guanine-Cytosine base pair
GC
15-25 ms
AT
5-10 ms
C
G
C
A A
G
*
*
4
1
23 4
1
A A
5
G
C
G
4
*
*
Base opening lifetimes
Base pair lifetimes (ms) 15°C
T
T
T
T
1 17 19 4
A A A A
A A A A
4 19 17
T
T
T
1
T
60 100 100 65 65 100 100 60
Leroy et al. Biochemistry
27, 1988, 8894
A4T4 versus T4A4
B-DNA - 2ns dynamic trajectory
Free energy calculations using
restrained opening
Guidice et al. ChemPhysChem 2, 2001, 673
Varnai & Lavery J. Am. Chem. Soc. 124, 2002, 7272
BIASED PROBABILITY HISTOGRAM
60
50
Nw
N(q)
40
 P*i(q) exp [Vi(q)]
30
i =1
20
Pi(q)  
10
0
Nw
FREE ENERGY PROFILE
30
 ni exp [Fi(q)Vi(q)]
i =1
W(q)
25
20
Nw
15
Fi(q)  kT ln  Pi(q)
10
i =1
5
0
Reaction coordinate (q)
WHAM
G A G A G A G A G A G A G
C T C T C T C T C T C T C
B-DNA oligonucleotide studied
Extraction d’une base de l’ADN
Closed AT pair
Adenine –50° (minor)
Adenine –100° (minor)
Adenine +50° (major)
Adenine +100° (major)
Free energy curves for base opening
T
G
Imino proton accessibility (Å2)
Dq Adenine (°)
Dq Thymine (°)
Dq Adenine (°)
Dq Thymine (°)
Base movements are coupled
T A-tract
T Ref
Dq
Sequence effects on opening: A-tracts
G
q < -50°
-50° < q < +50°
q > +50°
T
Bending amplitude (°)
A word of warning!
BASE FLIPPING
Hha1 methyltransferase
Klimašauskas et al. Cell 76 (1994) 357
Minor groove  Major groove
+160° opening
-200° opening
Backbone rearrangements
Backbone rearrangements
SUPERCOILING
DNA supercoiling (circular plasmid)
L = linking number = number of strand crossings
T = twist = number of turns of double helix
W = writhe = number of helix crossovers
L=T+W
s = supercoiling density = (L – L0) / L0 = DL / L0
typically s ~ -0.06 (1 crossing less per 17 turns)
DNA supercoiling
Linking number (L or Lk)
– a topological constant
Low force
Twist (T) versus Writhe (W)
High force
L=T+W
L.H.
R.H.
Interwound and toroidal forms of a
negatively supercoiled plasmid
Ethidium bromide intercalates into DNA
and reduces its twist by ~26°
Effect of an intercalator on a negatively
supercoiled plasmid
Topoisomerase I
- single strand cuts
- releases negative supercoiling
Topoisomerase II
(eukaryotes)
- double strand cuts
- releases negative supercoiling
Topo II (gyrase)
(prokaryotes)
- generates negative supercoiling
- consumes ATP
Reverse gyrase
(thermophiles)
- generates positive supercoiling
Topoisomerases
Topoisomerase I – single strand cuts
Topoisomerase II – double strand cuts
Topo II (gyrase)
DNA wrapping
DNA packed on nucleosomes
Nucleosome – schematic view
EXTREME DEFORMATIONS
DNA stretching
Cluzel et al. Science 271, 1996, 792
70 pN phase transition
S-DNA: fibre and ribbon forms
Greenall et al.
J. Mol. Biol. 2001, 305, 669
Rise ~ 5.6 Å
Helix spacing ~ 13 Å
Fibre diffraction of stretched DNA
TBP-DNA complex
5'
3'
3'
Minor
3'
5'
Major
5'
3'
5'
DNA: local stretching
DNA: global and local 3'3' stretching
X-ray
Model
TBP induced deformation
N
S
Biotin : Streptavidin
DIG : AntiDIG
Magnetic twisting control
Strick et al. Biophys. J. 74, 1998, 2016
Allemand et al. Proc. Natl. Acad. Sci. (USA) 95, 1998, 14152
Twisted DNA forms plectonemes
> 3 pN
< 0.3 pN
DNA twisting under tension
Simulation of DNA twisting
Simulating twisting