It is a well-known fact that double-stranded DNA forms
a double helix structure, but the double helix itself can
twist and turn into different conformations.
So, how does DNA change its shape,
and why is this behavior important
to our cellular functions?
This video will use a physical model of DNA
to help explain the significance of DNA topology
and simulate how specific enzymes modify the conformation
of double-stranded DNA.
The unstrained, or relaxed, DNA structure
has 10.4 base pairs per turn of the helix.
One way to describe the structure of a DNA double helix
is by a combination of its twist and writhe, the sum of which
is called the linking number.
The twist is described as the number of helical turns of one
strand about the other and can be measured
by counting how many times one strand wraps completely
around the other.
For example, this relaxed linear DNA
has a twist of one, two, three.
There is 0 writhe in this example.
The writhe is described as how many times the double helix
crosses over itself and is positive or negative
depending on the orientation.
For example, this circular DNA has a writhe of -1
because it follows the right hand
rule and the upper strand travels left to right.
And this DNA has a writhe of 1.
Both of these examples also have twist.
Realistically, long DNA molecules like these
can have linking numbers in the thousands.
For this video, we're going to look
at a physical model of circular DNA
with covalently linked ends.
Unlike linear DNA, this shape is topologically constrained,
meaning you can't change the linking number of the DNA
without breaking one or both of the strands.
But what would physically happen to this DNA
if you added or removed twists or writhe in the double helix?
Overwound DNA has fewer than 10.4 base pairs
per turn of the helix.
Thus, overwound DNA is more tightly wound than relaxed DNA
and has a greater number of twists
and a higher linking number than relaxed DNA.
When DNA is overwound and has an increased linking number,
the DNA becomes positively supercoiled,
and the double-stranded structure
begins wrapping around itself and creating positive writhe.
Regardless of these visible changes in conformation,
remember that positively supercoiled DNA
will have a higher linking number than relaxed DNA,
and the two strands of the double-stranded DNA
are more difficult to separate.
Underwound DNA has greater than 10.4 base pairs
per turn of the helix.
Thus, underwound DNA is less tightly wound than relaxed DNA
and has a fewer number of twists and a lower linking
number than relaxed DNA.
When DNA is underwound and has a decreased linking number,
the DNA becomes negatively supercoiled
and the double-stranded structure begins
wrapping around itself in the opposite direction
as positive supercoiling, creating negative writhe.
Regardless of these visible changes in conformation,
remember that negatively supercoiled DNA
will have a lower linking number than relaxed DNA,
and the two strands of the double-strand DNA
can be separated more easily.
But if DNA is least strained in the relaxed form,
how does the double helix change from being relaxed to overwound
or underwound?
And how does positively or negatively
supercoiled DNA return to a relaxed state?
Topoisomerases are a class of enzyme
that usually restores DNA to its relaxed state.
This means that topoisomerases unwind overwound DNA strands,
reducing the linking number back to the preferred 10.4 base
pairs per turn, or rewind underwound DNA strands,
increasing the linking number back
to 10.4 base pairs per turn.
Topoisomerases work by breaking one or two DNA strands
and passing the same number of DNA strands through the break.
This change results in an increase or decrease
in the linking number of our topologically constrained
circular DNA.
There are two main categories of topoisomerase, Type I
and Type II.
Type I topoisomerase breaks one of the two DNA strands
and passes the other strand through the gap.
This increases or decreases the linking number by 1.
Here, the top DNA double helix represents the structure
before the topoisomerase added a twist,
and the bottom DNA double helix is after the enzyme completes
the reaction.
Type I topoisomerase reactions do not
require additional energy.
Type II topoisomerase breaks both of the two DNA strands
and passes the entire double helix through the gap.
This increases or decreases the linking number by 2.
Here, the top DNA double helix represents the structure
before the topoisomerase acts, and the bottom DNA double helix
is after the enzyme completes the reaction.
In this case, the result is that the writhe of the molecule
increases by 2.
To make this idea clear, this diagram
shows what this change would look
like if you added one positive writhe at a time
instead of using Type II topoisomerase.
Type II topoisomerase reactions require some form of energy,
such as ATP or NADH.
Bacteria also have a special form of Type II topoisomerases
called gyrase.
Gyrase enzymes use energy to negatively supercoil
DNA rather than restoring DNA to the relaxed state.
Certain thermophiles also have topoisomerases
that use energy to positively supercoil
DNA rather than restoring DNA to the relaxed state.
Because it acts oppositely to gyrase,
these enzymes are called reverse gyrase.
But if DNA is unstrained in its relaxed state,
why would organisms have enzymes that intentionally supercoil
their DNA either positively or negatively?
Thermophiles and hyperthermophiles
thrive at relatively high temperatures,
ranging from 45 to over 100 degrees Celsius.
Positive supercoiling in thermophile DNA
adds additional twists or writhe to the double helix,
thus increasing the linking number.
These changes stabilize the structure,
preventing the DNA from denaturing
as easily at high temperature.
Wanting stable DNA structures makes evolutionary sense,
but what about the organisms with negatively
supercoiled DNA?
In fact, the DNA of most organisms
is negatively supercoiled.
Negatively supercoiled DNA provides a store of free energy
that helps with cellular processes that
require strand separation of the double helix,
like DNA replication and transcription.
Underwound DNA has a tendency to partially separate,
so strand separation is easier than in a relaxed DNA
structure, with more twists holding the double helix
together.
In short, negatively supercoiled, or underwound,
DNA makes it easier to separate the double helix
into two single strands.
As you separate the double helix and negatively supercoiled DNA,
you are creating more twists in the rest of the DNA,
causing rewinding of the underwound strands.
Rather than putting strain on the DNA topology,
the DNA that is still base paired
is returned to the ideal relaxed state.
Plus, you have a portion of separated single DNA strands
that can be used in cellular processes like replication
and transcription.
If you tried to separate the double helix of relaxed DNA,
you would introduce more twists in the DNA
and end up overwinding, or positively supercoiling,
the double helix, which is energetically unfavorable.
This video modeled how DNA topology
extends beyond the double helix structure
and how topology is critical to our cell's ability
to function smoothly by replicating or transcribing DNA
more efficiently.
Now, are you able to model overwound and underwound DNA
and tell the difference between Type I and Type II
topoisomerases and their functions?
And can you explain what role DNA topology
plays in cellular processes and function?
Thanks for watching.