Analysis of effects of Mechanical

Force on Bio-polymers
Rupture- Shearing of double stranded DNA

By- Tushar Modi

5th year Engineering Physics
M.Tech IMD
IIT(BHU)

Biopolymers = Bio + Polymers

These are the biologically occurring polymers in nature

Polymers are macromolecules made of certain basic units or
segments called monomers
A monomer itself can be anything from a simple molecule
consisting of a few atoms to a large complex molecule
These polymers are broadly of two kinds:
Homopolymers
Hetropolymers
Biopolymers are classified in three groups based on their
monomers:
Polysaccharides: made of sugars Eg. Amylose
Proteins: made of amino acids, Eg. Myoglobin
Nucleic acids: made of nucleotides, Eg. DNA, RNA

Importance of studying biopolymers:

They are involved in a number of crucial biological reactions.

They have a complex shape and a precise structure taking
part in different reactions. E.g. transcription (during cell
division), metabolism, signaling network in cells, etc.
Hottest topic for R&D is: Drug delivery and genetics.
Drugs are analogous to enzymes. They attach themselves to specific
locations or mix in system, and have some effect. Their delivery system
effects the time and intensity of action

Genetic: Study of Genes. They are the molecular unit of

heredity. They correspond to regions within DNA.
Their studies have proved to be very fruitful in designing of drugs and
genetic engineering. E.g. Marker: this is the gene or sequence of genes
on chromosome which is used to identify some specific trait.

Understanding of above reactions and processes, involve

understanding of the related biopolymers- structure, shape,
composition and kind of neighboring interactions

Single Molecular Force Spectroscopy (SMFS)

Single Molecule Experiments:
It is an experiment that investigate the properties of individual
molecules in a compound structure. They are different from
bulk experiments as in them the individual behavior of
molecules cannot be distinguished, and only average
characteristics could be measured.
They involve targeting some specific molecules and attach
probes to them. Then they are monitored and acted upon by
force and their motion is recorded to study dynamics of the
structure. These can be done my attaching magnetic beads to
molecules or by using atomic force microscope. Such
experiments have resulted in measurements of inter- and intramolecular forces in biological systems and have been able to
separate out the fluctuations of individual molecules from the
ensemble average behavior.

SMFS studies have provided unexpected insights into the

strength of the forces driving biological processes and
biological interactions responsible for the mechanical stability
of biological structures.
Surprisingly, with increasing number of experiments and
insights gathered so far, most of the interactions have a
physical and chemical nature and not just mechanical.
Several SMFS experiments these days are focused on free
energy landscape (FEL) of biomolecules; the protein
mechanical stability, experimental aspects of refolding under
quenched force , advances in optical tweezers etc.
Here the concentration is on the mechanical stability of dsDNA (double stranded) on application of a pulling force. It
can be in a direction parallel to the DNA axis (Shearing) and
perpendicular to the DNA axis (unzipping), rupturing and
twisting.

Some approaches for Biopolymer force analysis:

Analytic and statistical models:

In analytic models, we incorporate all the types of interactions
involved in the system in Hamiltonian.
The Hamiltonian is then used to find the equilibrium states of
the system and its properties.
However there are limitations to this approach as in here we
have to rely on a lot of assumptions in order to simplify the
calculations
A huge progress has been achieved using this approach in the
field of thermal denaturation of proteins, DNA, etc.

Coarse grain Simulations:

In this approach, an actual structure of the polymer is
constructed using the interactions discussed in Hamiltonian and
then we apply the external stimuli in the form of temperature
change or mechanical force or some chemical interaction.
In such a simulations, we dont have to worry about the
complexity of the solution as it is handled by computation and
thus a lot of assumption can be discarded giving us very accurate
results.

All Atomic Simulations :

These are the most accurate type of simulations possible.
In these kind of simulations, individual atomic interactions are
actually emulated by considering their potential (both classically
and quantum mechanically as the need arises). These atomic
models are then substituted to their actual positions in the
structure as it is calculated from experimental results.

Once the structure is made, any kind of external stimuli can be

applied over it in order to analyze the outcome. These
simulations are most accurate and almost dont require any
assumption regarding the structure. Effects due to solvent
molecules, molecular crowding or even twisting or distortions
can be incorporated.
However, these simulations are pretty costly to perform and
consumes a lot of time. These are usually done using a software
package for molecular dynamics called AMBER (Assisted Model
Building with Energy Refinement)

DNA Rupture

A DNA can be ruptured in three ways:

a) DNA unzipping
b) DNA rupture
c) DNA Shear
Source: Wikipedia

In my study, I have concentrated upon, the analytic models of

DNA shearing.

SHEARING OF ds-DNA
Model proposed
A double stranded DNA consists of two nucleotide polymer chains
joined together by hydrogen bonding bases. These nucleotide bases
are joined together by covalent bonds.
The interactions among the bases pairs are taken to be of
harmonic nature, and thus covalent bonds can be simply replaced
with harmonic springs. This assumption can be justified as under a
stretching force, the covalent bonds are stretched and thus the
entropic contributions are comparatively lower.
The hydrogen bonds potential can be simulated in a number of
ways like- harmonic (similar to covalent bonds), Lenard-Jones
potential or Morse potential. All these potentials give similar results
under the forces discusses here hence harmonic potential is used
because of its simplicity.
As the chain is stretched out, there is no need to take the
anharmonicity of the covalent bonds into consideration. It is also
cross checked with other methods, that even if we take them into
consideration, it will give similar results.

Another major assumption is taken in the formulation of

expression for the Hamiltonian. The diameter of DNA, i.e. the
space between the two strands are considered to be very small as
compared to the length of the polymer. Hence, the component of
force transverse to the direction of application of force can be
neglected.

Thus keeping the mentioned assumptions and formulations in mind,

following model can be proposed:
In the given figure, un represents the displacements in the upper strand having
a spring constant of Q, and U is the spring constant of lower strand where
displacements are represented by vn

The Hamiltonian of this system can be written as:

Results:
The Hamiltonian is differentiated with respect to displacement to find
the condition for minimum energy. The study of Hamiltonian gives
the following results regarding n = un vn (the elongation produced
in the nth Hydrogen bond.
The hydrogen bonds between the strands, elongate under the force.
Their elongations are given as:

where, A and and are the constants depending upon the system
configuration given as:

and,

The other constant 0 is the elongation of the central hydrogen bond

and its value for a particular force can be calculated using boundary
conditions for conservation of force.

Plot between the Displacements produced in hydrogen bonds in base pairs

with their indices

The force acting being exerted on each base pair is given as =

(displacement produced) x (spring constant) (Hookes law). Thus the
net shear force couple acting on the polymer can be given as the
summation of the force exerted by all the base pairs:

The shearing starts from the point of maximum elongation, which can
be seen from the elongation equation and applying the required
conditions the Force required to shear off the whole DNA chain is
given by :

where the constant f1 is the force required to rupture a single

hydrogen bond; and l is the total number of base pairs in the DNA.

Plot of the critical shear force with total number of base pairs

Analysis:

The plot between elongations produced in hydrogen bonding

and their indices in the DNA chain was an asymmetric one.
Thus for an applied force, one end will be more stretched
than the other one and hence clearly the shearing will start
from a particular end regardless of the direction in which the
force is applied. It was a ground breaking finding which
explained why the genetic code is preserved in offspring even
when DNA shearing and transcription happen in a very
random fashion during a cell division.
The force required to shear off a DNA, as seen in the plot
between critical force and the total length of the polymer
chain, at first increases linearly with the length and then
reaches a saturation limit after which increase in length has
no effect. Similar results were obtained in experimental
studies of the same. Thus howsoever long DNA sequence is
present in cells it only requires a finite amount of force to
open it up.

Discussion:
The model presented was remarkable because of its simplicity with
a good prediction of properties, however a lot needs to be done.
The model has a major limitation that it does not include the effect
of temperature. There is no scope to include thermal noise
fluctuations and effect of environmental effects (like solvent and
molecular crowding) in the formulations.
We have gone for a publication for this study and have received a
good review from Journal of Chemical Physics and are on a fast
track to get a final publication.
Apart from the study of shearing, a lot of analytical work as well as
statistical formulations are also done in unzipping transitions of
DNA where we have used transfer matrix method to solve
integrals numerically making C programs for the same. Their results
have also shown remarkable similarity with other all atomic as well
as experimental studies giving us a very a promising prospective in
research for the same.
Currently I am involved in making analytical formulations for
shearing taking heterogeneity, temperature effects and structural
complexities like helical structure, randomness, etc. into
consideration

Thank You