Terrm 2 Lecture 15: The Mechanical Properties Of DNA Can Encode Genetic Info (Aakash Basu)

Question 1

Protein binding deforms DNA, mechanical distortion can be measured

Answer 1

Mechanical distortions require cell energy to be carried out. Can be measured by tethering a length of DNA to a surface and attaching the other end to a plastic bead. This bead can be held in an optical trap (like a focused laser beam) that forces the bead to stay in the middle of the beam.
Moving the beam forward/backwards pushes and pulls on the bead which in turn stretches the DNA and the force can be measured by force (pN) against extension (nm). DNA shorter than 150bp is very hard to bend.

Question 2

When proteins interact with DNA they tend to interact with lengths 10-50bp or 100bp at most

Answer 2

At this length DNA is very rigid
Flexibility also depends on local sequences so energetic cost associated with deforming DNA depends on local sequences so genomes can mechanically control and regulate DNA protein interactions

Question 3

Basu et al 2000 looked into bendability of short DNA fragments

Answer 3

Using fragments (100bp) with short complementary ss overhangs that allowed them to hybridise and bp to form loops.
Flexible DNA forms loops easily while rigid DNA will take longer.
They measured how quickly linear DNA formed loops to give a measure of bendability for the DNA fragments tested.
Fluorescence resonance energy transfer (FRET) is used to measure this Fluorophores hit with a certain light wavelength absorb a proton and emit the photon at a slightly lower wavelength
The method used 2 Fluorophores one at each end of the DNA fragments. One that absorbs the same wavelength as the other one emits. When far apart the two cannot interact and when close together one can excite the other.
So it is possible to tell if the DNA is linear or loop and track the rate of loop formation over time.
In theory eventually all loop eventually but some take longer than others as speed related to flexibility.
Measuring changes in local flexibility across a whole genome is possible by chopping it all up into 100bp lengths of overlapping fragments.
Originally this process was very slow so modern technology has been developed to measure hundreds of thousands of fragments rapidly

Question 4

The region just upstream of a start site must be without nucleosomes

Answer 4

This allows DNA Pol to bind to this region known as a ‘nucleosome depleted region’ this depleted region is inglexible (Basu et al 2021)

Local flexible peaks occur where nucleosomes form

Question 5

Conclusions

Answer 5

• we can make physical measurements of the properties of DNA
• sequence can encode the local physical properties of the DNA polymer
• it is therefore possible for genomes to use sequences to control physical properties of DNA and impact various biological processes
• several mechanical features of yeast are conserved in fly and mouse genomes
• DNA mechanisms are selected for by evolution