DNA Super Coiling:
Whether the DNA is single stranded or double stranded, inside the cell it is always subjected to bending, folding, over winding and under winding. Such changes make stable DNA into unstable or vice versa, because energy constraints. Thermodynamically complaint B- DNA contains 10.6 bp per turn, and such DNA without any constraints or stress or torsion, when placed on a flat surface it lies flat as a circular molecule or a linear molecule. Such DNA, which is free from stress, is called Relaxed DNA.
This is an EM of a Super coiled and one relaxed DNA
When a relaxed DNA is subjected to bends, or openings of DNA, over winding or unwinding, its base pairs per turn changes, and the DNA is subjected to stress and strain. In order to overcome such distortion, which has rendered the DNA unstable, the DNA twist around itself, like a circular rubber band undergoing twisting, such a twists on its own thread is called Super coiling; it is also referred to as tertiary structure.
To explain super coiling phenomenon, if a 5300 bp long circular DNA, which contains 500 helical turns with 10.6bp per turn, is twisted 50 times by 360^0 each time, in right handed direction i.e. towards its own right handed coil, the DNA becomes over coiled or over winded and the helix becomes tight. It virtually means adding 50 more helical coils to the already existing 500 helical turns, making the total number of turns to 550. So the 5300 bp long DNA has to accommodate 50 more turns, which is torsionally a stressed condition. If 550 divide 5300 bp, the number of base pairs per turn will be 9.6; it is a drastic deviation from the normal and thermodynamically feasible and stable state of 10.6 per turn. This over winding is thermodynamically incompatible, so in order to accommodate the excess number of coils, the helical DNA twists on its own length in space in left handed direction, like a rubber band twist, in left handed way, such a twist is called super coil; in this case the super coiling is positive. Such super coilings overcome stress energy and accommodate the extra 50 numbers of added coils to the existing 500 helical turns. It is because of super coiling, now the number of base pairs per turn would be 10.6
Negative super coiling:
Similarly, if a 5300bp long circular DNA with 500 turn is twisted 50 times 360^0 each, in left handed way, the DNA gets under wounded, and likely the DNA strands can separate. In this case the number of turns lost are 50 and to adjust the number of coils, the number of base pairs per turn will be11.6, which is not thermodynamically feasible, hence the DNA takes a right handed twist on its own; the DNA helix is in the form of negatively super coiled DNA. This super coiling accommodates the stress energy and the DNA becomes stable with 10.6 bp per coil. Generally the density of the super coils a DNA is one super coil per 200bp or per ~20 turns of the helix. Negative super coils help in stabilizing certain DNA structures i.e. Z-DNA, cruciform DNA, triplex DNA. It also helps in unwinding of the DNA during replication as replication bubble or during transcriptional initiation as transcriptional bubble.
Detection of super Coils:
Super coiled DNA can be distinguished from the normal DNA using gel electrophoresis. Super coiled DNA moves faster in an agarose gel than the relaxed DNA. If a purified plasmid DNA is run on the gel, and if one finds more than two bands, it means the one that moved faster is fully super coiled and the one that moves slower is nicked, so it moves slow. By subjecting such super coiled DNA to Topoisomerases, an enzyme that nicks and relaxes the super coiled DNA, one cut per digestion and such digested DNA if ran on an agarose gel, one can observe a number of bands from the bottom. The bottom most is the most super coiled and uncut. The band found next to the uncut shows that it is relaxed by one super coil. The uppermost band is the one, which is totally relaxed. The number of bands found on the gel gives the total number of super coils in a given DNA.
During replication, transcription, recombination, DNA repair and bending or folding of DNA or during gene expression or compacting of DNA as chromosomes, the normal relaxed DNA of 10.6 bp per turn, dramatically changes its base pairs per turn and the DNA is subjected to torsional stress; to accommodate such strain the DNA has to undergo super coiled states. Such topological changes in DNA can described by three parameters, they are
Linking number, twist and Writhe.
Linking number (L):
It is an invariant but an integer value. It is the sum of two geometric components, one twist and the other writhe. Twist refers to the number of base pairs per one helical turn.
The number of helical turns in a given DNA molecule of certain length, say in this case the DNA is double stranded, circular and 5300 bp. The number of twists = base pairs per turn or the helix. In a relaxed DNA, in this case, 10.6 bp/ turn, it amounts to 500 helical turns.
It is super coil (SC) that twist on its own length like a rubber band twist. The helix crosses over it in space., the writhe can be plectonic or toroidal.
L = T + W;
When L < T + W, the DNA is in (-) SC state,
When L > T + W, the DNA is in (+) SC state,
L = T + W, where W = 0. Relaxed DNA;
Ex. DNA is ds, circular, 5300 bp long, bp/turn= 10.6, number of turns (360^0) = 500, Sc or writhe is Zero.
L = T + W;
Delta L = dT + dW,
Under wound= -dL = (-) SC
Over wound = +dL = (+) SC
Change in the state of DNA can be described by specific differences in linking numbers, i.e. D = dL / L (0),
So the L is; L = 500 + 0; so L = 500 turns, it also is also denoted as L (0)
If one wants to change the linking number, it is possible to do so, by nicking one of the strands, introduce certain number of turns, either way, then ligate the ends. The number of SC and handedness of the SC depends on the direction of turns and the number of turns. Let us take an example to explain this
Negative super coils:
The DNA is 5300 bp,
The number of left handed turns (360^0) introduced is 50 that mean the number of turns removed is 50 = 530 bp.
Calculate the number of writhes. In order to accommodate the loss of number of coils, it has to accommodate11.6 bp per turn, but energetically it is unfavorable, so it produces 50 negative super coils so as to have 10.6 bp/turn as an unstrained molecule. The 50 negative super coils are 180^0 each and strands crossing over is on right handed direction.
T + W500 (5300
Positive super coils:
The DNA is 5300 bp, 10.6bp/turn, so T=500 turns
The DNA is tuned right handed 50 times 360 ^0 each.
So the number of turns is 500+50=550, if this has to be accommodated the number of base pairs per turn will be=9.6; 5300%550 = 9.6, this bp is energy wise not compatible, so the DNA produces 50 positive super coils or writhes to be compatible. Each super coil is 180^0 and the DNA crossing is left-handed.
And L > L0
Generally, most of the times, positive super coils develop, when DNA opens and replication progresses in both directions; transcription also causes positive super coils. When DNA winds around histone complex to form Nucleosomes DNA experiences negative super coil state. Binding of proteins or protein complexes to DNA either during Replication or recombination or transcription, DNA is subjected to such stress and strains. Positive coils act as hindrance to the progress of replication fork as well as transcriptional fork and such positive super coils are removed and DNA is either rendered negative super coils or relaxed, which is essential for the said functions, otherwise DNA fails to replicate and fails to transcribe, which has serious consequences to the cell. Even ssDNA, such as circular DNA are in super coiled, with out which the DNA fails to replicate.
Opening of DNA causes over winding of DNA helix ahead of the replication fork,i.e positive super coiling
This diagram just exhibit how thw DNA can be knotted into different structural forms
The figure (a) shows how a DNA can be supercoiled and (b) represent catenation of DNAs
This Diagram is #-D model of SC DNA
Topoisomerase II in
This a linear relaxed DNA
This is relaxed circular DNA
The Topoisomerase has both nicking and ligase activity, here it shows ligase activity
Topoisomerase II in action, where two strands are cut and swiveled into each other
Topoisomerase Ribbon Model