DNA: Structure, Dynamics and Recognition

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DNA Structure, Dynamics and Recognition
L4 DNA deformation
Les Houches 2004
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BASE PAIR OPENING
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Biological time scale
Bond vibrations 1 fs (10-15 s) Sugar
repuckering 1 ps (10-12 s) DNA bending 1
ns (10-9 s) Domain movement 1 ?s (10-6 s) Base
pair opening 1 ms (10-3 s) Transcription 2.5 ms
/ nucleotide Protein synthesis 6.5 ms / amino
acid Protein folding 10 s RNA lifetime 300 s
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Enzymatic base chemistry
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Adenine-Thymine base pair
HN3 imino proton
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Guanine-Cytosine base pair
HN1 imino proton
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Base opening lifetimes
GC 15-25 ms AT 5-10 ms
C G C A A G A A G C
G 4 1 1 23 4 5 4

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A4T4 versus T4A4
Base pair lifetimes (ms) 15C
T T T T A A A A 1 17 19
4 4 19 17 1 A A A A T
T T T 60 100 100 65 65 100 100 60
Leroy et al. Biochemistry 27, 1988, 8894
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B-DNA - 2ns dynamic trajectory
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Free energy calculations using restrained opening
Guidice et al. ChemPhysChem 2, 2001, 673 Varnai
Lavery J. Am. Chem. Soc. 124, 2002, 7272
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WHAM
BIASED PROBABILITY HISTOGRAM
Nw
? Pi(q) exp ??Vi(q) i
1 Pi(q) ? ?????????? Nw
? ni exp ?Fi(q)?Vi(q)
i 1
Nw Fi(q) ? ?kT ln
? Pi(q) i 1
N(q)
FREE ENERGY PROFILE
W(q)
Reaction coordinate (q)
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B-DNA oligonucleotide studied
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Extraction dune base de lADN
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Closed AT pair
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Adenine 50 (minor)
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Adenine 100 (minor)
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Adenine 50 (major)
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Adenine 100 (major)
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Free energy curves for base opening
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Imino proton accessibility (Å2)
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Base movements are coupled
Dq Thymine ()
Dq Adenine ()
Dq Adenine ()
Dq Thymine ()
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Sequence effects on opening A-tracts
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Bending amplitude ()
q lt -50 -50 lt q lt 50 q
gt 50
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A word of warning!
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BASE FLIPPING
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Hha1 methyltransferaseKlimaauskas et al. Cell
76 (1994) 357
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Minor groove ?? Major groove
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-200 opening
160 opening
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Backbone rearrangements
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Backbone rearrangements
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SUPERCOILING
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DNA supercoiling (circular plasmid)
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DNA supercoiling
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)
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Linking number (L or Lk) a topological constant
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Twist (T) versus Writhe (W)
Low force
High force
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L T W
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Interwound and toroidal forms of a negatively
supercoiled plasmid
L.H.
R.H.
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Ethidium bromide intercalates into DNA and
reduces its twist by 26
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Effect of an intercalator on a negatively
supercoiled plasmid
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Topoisomerases
Topoisomerase I - single strand cuts - releases
negative supercoiling Topoisomerase II - double
strand cuts (eukaryotes) - releases negative
supercoiling Topo II (gyrase) - generates
negative supercoiling (prokaryotes) - consumes
ATP Reverse gyrase - generates positive
supercoiling (thermophiles)
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Topoisomerase I single strand cuts
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Topoisomerase II double strand cuts
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Topo II (gyrase)DNA wrapping
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DNA packed on nucleosomes
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Nucleosome schematic view
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EXTREME DEFORMATIONS
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DNA stretching
Cluzel et al. Science 271, 1996, 792
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70 pN phase transition
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S-DNA fibre and ribbon forms
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Fibre diffraction of stretched DNA
Greenall et al. J. Mol. Biol. 2001, 305,
669 Rise 5.6 Å Helix spacing 13 Å
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TBP-DNA complex
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DNA local stretching
5'
3'
3'
Minor
3'
5'
Major
5'
3'
5'
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DNA global and local 3'3' stretching
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TBP induced deformation
X-ray Model
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Magnetic twisting control
N
S
Biotin Streptavidin
DIG AntiDIG
Strick et al. Biophys. J. 74, 1998, 2016 Allemand
et al. Proc. Natl. Acad. Sci. (USA) 95, 1998,
14152
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Twisted DNA forms plectonemes
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DNA twisting under tension
gt 3 pN
lt 0.3 pN
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Simulation of DNA twisting
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Simulating twisting