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INTRODUCTION TO
Lecture 1
KEY EXPERIMENTS ON
Topo II DNA preferences DNA knots Topo II mechanics DNA relaxation in vivo
Lecture 3 Lecture 2
(+)
?
Eukaryotic TOPO II G::T Bacterial GYRASE G:T
(-)
Bacterial TOPO IV
Roca (2003)
Does TOPO II
NO
MONOMER circle
LIG
DIMER circle
NO
!!!
DECATENATION :
Roca (2002)
DNA CATENATION :
DNA KNOTTING :
Roca (2002)
With ( -- ) supercoiled, TOPO II is more prone to decatenate With ( + ) supercoiled, TOPO II is more prone to catenate
WHY ?
Yes
Roca (2002)
( + ) & ( -- ) SUPERCOILS are EQUALLY removed ( -- ) (+) SUPERCOILS favour DECATENATION SUPERCOILS favour CATENATION
Roca (2002) Trigueros et al. (2004)
(R)
( -- )
TRIPLE NODES may ORIENTATE the TOPOISOMERASE TO UNLINK DOMAINS without RELAXING SUPERCOILING
DNA TOPOLOGY
INTRODUCTION TO
Lecture 1
KEY EXPERIMENTS ON
Lecture 3
DNA Knots
Topoisomerases Recombinases
nick
nick
+ +
41
+ 31
52
51
+3#-3
61
62
63
mirror
10 3 bp 10 4 bp 10 5 bp
>
> 96 % 2%
>
< 0.2 %
Rybenkov, V. V., Cozzarelli, N. R., and Vologodskii, A. V. (1993). Probability of DNA Knotting and the Effective Diameter of the DNA Double Helix. Proc. Natl. Acad. Sci. USA. 90, 5037-5311.
In 1982, DNA extracted from a bacterial virus (phage P4) was found to be cyclisated and highly knotted
Knotting probability ~ 1
P4 genome is a 10 kb linear DNA molecule with cos ends DNA cyclisation normally occurs after DNA is injected inside the bacteria
DNA
Unknotting by TOPO II
0 1 2 3 4
min
Arsuaga, J., Vazquez, M., Tigueros, S., Sumners, D. W., & Roca, J. (2002) Knotting probability of DNA molecules confined in restricted volumes: DNA knotting in phage capsids. Proc. Natl. Acad. Sci. USA 99, 5373-5377.
Computer simulations of random polygons confined to spherical volumes demonstrate that Knotting probability and Knot complexity increase by the effect of confinement
Arsuaga et al (2002)
KNOT SPECTRA
EM by Stasiak
+ 41
+
Wr = 0.00
+ + +
51R
+
Wr ~ + 6.26
Computed distributions of the knots 41, 51, 52 in ideal chains randomly confined to a sphere of fixed radius, when conformations with writhe below a given value are rejected.
Relative probability
N = 90
R=4
simulation
51
52
41
0 2 4 6 8
104 bp (~ 3
m)
108 bp (~ 30 cm) 10 m
40 nm