INDIAN INSTITUTE OF TECHNOLOGY ROORKEE

NPTEL

NPTEL ONLINE CERTIFICATION COURSE

Biomedical Nanotechnology

Lec - 09
Bio-Nanomachines

Dr. P. Gopinath
Department of Biotechnology
Indian Institute of Technology Roorkee

Hello everyone, I welcome you all to the 9th lecture of this course. This 9th lecture is on bio nano
machines.
(Refer Slide Time: 00:28)

In this lecture, we will learn about DNA and protein based nano machines. I will also
demonstrate a simple experiment to study the motility of bacteria. We will also learn about the
communication of nano machines.
(Refer Slide Time: 00:39)
First we will see what nano machine is. Nano machines are nano sized gadgets, doing
mechanical work. Examples are nano gears and nano wheels. How can we make nano motors?
What are the approaches available for making nano motors?
(Refer Slide Time: 00:55)

There are three approaches for making nano motors. They are physics, chemistry and biology
based methods. In physics based approach, we can use carbon nano tubes and make nano
machines or nano motors. In the chemistry based approach, we can use the supra molecular
chemistry and make small nano sized machines and nano sized switches. In the biology based
approach, we can use DNA and proteins for making various nano sized machines or rotors.
(Refer Slide Time: 01:24)

How to construct this nano machine? The first step for constructing nano machine is modeling
and designing, which comes under nanoarchitectonics. For synthesis of nano materials, mainly
chemistry and bio-chemistry contributes. The next step is fabrication. For fabrication of nano
devices, we need the contribution from cell biology and other fields. Finally we have to integrate
and make the complete nano machine. So what would be the application of this nano machines?
Let us see an example.
(Refer Slide Time: 02:05)
We can make a drug delivery nano bot, that is, nano robots which can precisely reach the tumor
location and release the anti-cancer drug. Here, the nano robots are specifically binding to the
cancer cells and releasing the drug. We can make such kinds of small sized nano machines not
only for drug delivery applications but also for various other applications.
(Refer Slide Time: 02:35)

For making nano machines, we can take the idea form the cells. Because the cell is the most
sophisticated existing nano machine and it has an efficient energy consumption. It also has multi-
tasking, computing and multi sensing capacity. So by mimicking cells, we can make such nano
scale machines or nano scale Robots.
(Refer Slide Time: 02:55)

Cells perform complex metabolism every milliseconds or nano seconds. These happens without
any disturbance or without any traffic inside this cell. By understanding this complex processing
inside the cells, we can make very small sized, nano scale machines, which can do the job very
precisely and which will have wide applications.
(Refer Slide Time: 03:23)
We can make biologically inspired nano machines.
(Refer Slide Time: 03:27)

There are two approaches for making nano machines- bottom up and top down approach. So man
made nano machines follow top down approach. We started with robots and we are trying make
nano robots. But nature works as bottom up approach. It assembles DNA and protein, and makes
the nano scale machines.
(Refer Slide Time: 03:50)
And especially in case of nano machines, the DNA and protein plays a major role because the
manipulation of DNA or hybridization will allow us to obtain user defined biological nano
machines. This means we can easily manipulate or modify the DNA and we can make such kind
of small nano machines. Further, we can also make user defined biological nano machines.
These are some of the advantages of DNA and protein based nano machines.
(Refer Slide Time: 04:20)
So in the previous lecture, I already told you that A always forms bond with T and G always
forms bond with C. I have also explained already why DNA is helical in nature and what the
reason for the helical structure of DNA is.
(Refer Slide Time: 04:35)

Now, let us see the various forms of DNA. We have 3 forms of DNA that is A DNA, B DNA
and Z DNA.
(Refer Slide Time: 04:55)
So the main difference is that A DNA has 11 base pairs per turn of helix, B DNA has 10 base
pairs per helix, and Z DNA have 12 base pairs per helix. So using this how to make nano
machines and nano motors?
(Refer Slide Time: 05:02)

Before we see how to make nano machines, let us get an idea about some of the terminologies
used. You can see here, these are double helical region and this b is a sticky end. That means
over hangs. C is your bulge loop and d is a hairpin loop. If you have this kind of DNA sequence,
it is called as hairpin loop. Further, this e is a junction and this f is a cross over.
(Refer Slide Time: 05:28)
We can stretch the DNA, condense the DNA and also we can play with the DNA by using
external fields.
(Refer Slide Time: 05:37)

Let us learn about protein synthesis now. We get the RNA from the DNA through the process of
transcription. From RNA, proteins are formed by translation. By taking the examples of
transcription and translation, we can make such kinds of small nano machines.
(Refer Slide Time: 05:56)
Let see how translation is done. This is the mRNA, this is the tRNA and this orange color is the
rRNA. The tRNA is coming and binding to mRNA. Another tRNA is coming and if it is not
matching with the sequence in the mRNA, it will be removed. When the sequence in the tRNA is
matching with the sequence in the mRNA, a peptide bond is formed. So similarly one by one the
amino acids will come and join to form the protein sequence. Hence, it is also a kind of nano
machine and at the end of the reaction the mRNA is degraded. Here, the poly A tail sequence in
the mRNA is very important. If the length of the poly A tail is more, that means the half-life of
the mRNA will be more. By mimicking this biological phenomenon, we can make nano
machines.
(Refer Slide Time: 06:59)
So let us see another example for how to make DNA based motors. We can use the DNA and
RNA polymerase enzyme. So what happens is, when the enzyme synthesizes the RNA, the DNA
will start rotating in this direction. We can easily measure this by tagging the DNA with the
magnetic bead. The top of the magnetic bead can be tagged with fluorescence beads. This
magnetic bead will be hold by external magnetic field.

When we add the RNA polymerase enzyme, RNA molecules will be made. When this process
happens, the DNA will rotate in this direction and this rotatory motion can be easily monitored
using the microscope.
(Refer Slide Time: 07:39)
So the next example is, DNA based nano machines. Various machines are available as DNA
based nano machines, like B-Z transition, molecular tweezers, PX and JX2. We will see them one
by one.

(Refer Slide Time: 07:54)

The first one is B-Z rotator. This machine is based on the transition between B-DNA and Z-DNA
by changing the ionic strength of the medium. This motion can easily be monitored by FRET.
What is FRET? It is a mechanism describing the energy transfer between two light sensitive
molecules. Here, we are simply changing the ionic condition of the medium.
(Refer Slide Time: 08:20)

We can convert the B-DNA into Z-DNA. It has a fluorescent molecule and a quencher. So when
it is in the B-DNA form, it would not show any fluorescence. But when the ionic condition is
changed, it will become a Z-DNA. Then the fluorescent molecule and the quencher will be
separated and it will give the fluorescence.
(Refer Slide Time: 08:43)
So when you change the ionic condition, this B-DNA will become Z-DNA and your
fluorescence and quencher will be separated, which gives fluorescence. This is called as B-Z
transition. So by simply changing the ionic condition from B-DNA we can make the Z-DNA.

In the B-DNA, the fluorescence and quenchers are together so it would not show any
fluorescence signal. But when it becomes Z-DNA, the fluorescent molecule and the quencher
will be released and so this will give the fluorescence signal.
(Refer Slide Time: 09:46)
The next example is rotatory DNA nano machine. This device works by producing two different
confirmations- parallel confirmation and zigzag confirmation by using two pairs of strands which
are called as set strands. This will bind to the single stranded DNA and it will make this kind of
parallel or zigzag confirmation. So here you can see the example of double stranded DNA
holding the single stranded DNA. When we add set strands, it will form this kind of parallel
confirmations and zigzag confirmations, depending on the size of the set strands. If the set
strands are matching with the size of the single stranded DNA, it will produce parallel
confirmation. If it is slightly bigger than the single stranded DNA, then it will form a zigzag
confirmation. So here, the DNA is acting like the fuel. Based on the DNA it will have a rotatory
motion.
(Refer Slide Time: 10:44)

Next example is walking triangle. It consists of a particular kind of a DNA sequence and when
another short fragment of a single stranded DNA is added, it binds with this single stranded
DNA. In between this two triangles, there are also some unbound DNA strands, which will form
a loop like structure. So in this case, both the triangles should be on the top and when we add the
red color DNA strand, which is matching with the blue color single stranded DNA. When it
forms a hybridization what happens is, this triangle will have 180 0 rotation and it will move to
the bottom. This is another example for a DNA based nano mechanical device.
(Refer Slide Time: 11:27)

Another example is DNA tweezers. Here, you can have a fluorophore that is the
tetrachlorofluorescein. In short it is TET and another one is quencher that is TAMRA
(tetramethylrhodamine).
(Refer Slide Time: 11:48)
So using these dyes, we can make the DNA tweezers. We can open the tweezers or we can close
the tweezers. We can also add a single stranded DNA. So this blue color will bind here and this
green color will bind here and it form this kind of structure. When it form this kind of structure,
the fluorescence and quencher are close together, which means there will be no fluorescence.

And when we add another DNA sequence that is highly specific for this blue color and green
color strands, it will combine and this strand will be removed. So when it is removed, the
structure will be in an open state. Then the fluorescence molecule and the quencher molecule are
separated and so it will give a fluorescence signal. So in this picture, you can see here that the
fluorescence intensity is more when it is open state. The fluorescence intensity is less when it is
in the closed state.
(Refer Slide Time: 12:50)

Now, let us see another example for a protein based nano machines, that is Dynein, F1ATPase
and bacteria flagella.
(Refer Slide Time: 13:00)
The Dynein is a molecular motor that walk along the microtubule in a cell. The ATPase is an
enzyme that synthesizes ATP by using an influx of protons to rotate. The bacteria uses the
flagella to move from one location to other location towards the chemical that is food.
(Refer Slide Time: 13:19)

First we will discuss about bacterial chemotaxis. The bacteria move from one location to another
location using the flagella. It generally move towards the food. It gets the signal from the food
and it moves toward that food. This phenomenon is called as chemotaxis because the bacteria get
the chemical signals and then it moves towards the food. Here, the bacteria move using the
flagellar motors.

This protein network directs movement based on the external conditions. If there is food that is
an attractant or any other chemical which may be a repellant. The presence of an attractant or a
repellant will simulate chemotaxis network. More than 7 proteins are activated and only then this
flagella will be in action to move towards the food. The chemical attractants bind to the
chemoreceptors and the chemoreceptors transmit information to a central processing system. The
central processing system integrates many inputs and sends the signal to control the flagellar
motors.
(Refer Slide Time: 14: 02)

So from this bacterial motility, we can take the idea of sensitivity and we can develop such kind
of nano machines and nano robots, which can sense the cancer cell in our body. Such machines
can reach the cancer location and deliver any anti-cancer drug.
(Refer Slide Time: 14:59)
Consider this example. We have sugar solution. All the bacteria are moving towards the sugar
solution. The bacteria swim by rotating the flagella and the motors located at the junction of the
flagellum. This motor can rotate clock wise or counter clockwise. The bacteria swims smoothly
for 1 second and then it changes the direction of motion by an average of 60°. Hence, the
movement is with respect to the attractants. If we have increasing concentrations, then the
motion will be uniform and be less tumbling. If we are having decreasing concentration of the
solution, the bacteria will have more tumbling motions.
(Refer Slide Time: 15:39)
By using the hanging drop method we can study the motility of bacteria. For this we need a
concave glass slide. This glass slide will be having a concavity. We also need a cover slip. On
the four sides, we will be adding Vaseline. In the middle of the cover slip, add 5 to 10 µl of
bacteria sample. In this example, we are going to add GFP E-coli, which is a green fluorescence
protein expressing E.coli. So by using this bacteria we can easily monitor the motility of bacteria.

Once we add these bacteria to this cover slip, then gently keep this glass slide on the top and
invert the slide. So when you invert the slide, the cover slip will be on the top of this concave
slide and your bacterial solution will be hanging.
(Refer Slide Time: 19:16)

That is why it is called as hanging drop method. Under the microscope, we can study the motility
of bacteria.
(Refer Slide Time: 19:26)
Now let us see how the ATP synthase works. Here the proton gradient drives the F1 rotation
which is accompanied by the synthesis of ATP from ADP.
(Refer Slide Time: 19:45)

If you have high ATP concentration, then the rotation will be in the opposite direction, resulting
in ATP hydrolysis. This action pumps the protons to outside. Hence, during ATP synthesis, the
rotation will be in clockwise and during ATP hydrolysis, it will rotate in the counter clockwise
rotation. Hence, it is a kind of a reversible motor.
(Refer Slide Time: 20:00)

Here the F1 unit is attached with the dye-loaded actin and attached to F0. We can easily see the
movement of gold bead by laser optical imaging. This is the gold bead. You can see the rotatory
motion. The motor takes three 120° steps to complete one rotation. It hydrolyses one ATP
molecule per step.
(Refer Slide Time: 20:28)
So now let us see a nano machine communication.
(Refer Slide Time: 20:33)

Why this nano machine communication is important? The nano machines such as the chemical
sensors like nano- valves and nano switches cannot execute the complete task by themselves. So
the exchange of information and commands between the networked nanomachnies will allow
them to work in a cooperative and synchronous manner. This facilitates to perform more
complex functions in the body such as drug delivery as well as disease treatment applications. So
the nanomachine communication is very important if you want to make a successful nano device
or nano motor for drug delivery applications.
(Refer Slide Time: 21:10)
Let us see biological nano machines. We can make different kinds of logic gates. Only if both
substrate and effector is present, the product will be made. If there is no effector, then the
substrate remains unchanged. So we can make different kinds of logical gates using this
principle. For example, if the cancer cells are expressing two kinds of receptors or two kinds of
markers, only then the nano particles will bind. When it binds based on this condition, it will
release the drug. This way, the nano machines will not release the drugs into any healthy cells.
By designing this kinds of logic gates, we can target the cells for various drug delivery
applications.
(Refer Slide Time: 21:44)
In this molecular communication network, both the sender and receiver are biological nano
machines. The communication carrier is also a molecule that is in the range of nano scale.
Proteins, ions and DNA are the communicating molecules and these are in the range of nano
scale only. The communication distance will be in nano or micro scale and the receiver which
receives the signal will also in the range of nano scale.
(Refer Slide Time: 22:07)
The sender is a nano machine and the receiver is also a nano machine. The information which is
going from sender to receiver is also in the nano scale that is, they are protein or ions or DNA.
(Refer Slide Time: 22:18)

Here, the molecular communication is divided into two types. The molecular motors that are
wired and the molecular motors based on calcium ions which are wireless. You can simply
assume that this molecular motor is similar to the telephone connection or landline connection
and the calcium ion based molecular motor is the wireless network or the cellular network.
(Refer Slide Time: 22:41)
So let us see how communication happens using molecular motors. First we will discuss about
what is a molecular motor. It is a protein complex that transforms chemical energy into
mechanical work at a molecular scale. It has the ability to move the molecules. The molecular
motors carry the information from one location to other location through the microtubules.
(Refer Slide Time: 23:06)
It is mainly found in the eukaryotic cells. It can carry the message and walk on the microtubules.
So the molecular motors travel or move along the molecular rails which are the microtubules.
The movement created by the molecular motors can be used to transport information molecules.
(Refer Slide Time: 23:24)

The nano machines communicate using the molecular motors and this molecules are transported
from encoder to decoder using this network of rail. It is this molecular motor, which will walk on
this network of rail and reach from encoder to decoder.
(Refer Slide Time: 23:46)
It will carry the information. It selects the right molecules that represent the right information
and it will carry the information through the microtubules. It moves and reaches the receptor.
Then the molecule is detached and it will perform the work according to the information carried.
(Refer Slide Time: 24:24)

The next example is communication using calcium signaling. Here two different deployment
scenarios are available. The first one is the direct access and another one is the indirect access. In
direct access, the exchange of information is among the cells located next to each other. In
indirect access, the cells can be separated without any physical contact and the information can
reach the receiver.
(Refer Slide Time: 24:38)

Now, let us see the direct access. All the cells are connected through gap junctions. There are
nano scale protein channels between two adjacent cells, which allow small molecules to be
shared among the cells. This enables all the coordinated actions of the cells.
(Refer Slide Time: 24:57)
In direct access, the calcium signal travels through the gates that is the gap junction. From here,
the information will go to the decoder and receiver through the gap junction of the cells.
(Refer Slide Time: 25:13)

Next one is indirect access. Here the nano machines release the information molecules to the
medium. So the calcium molecules can go to the receiver which is little far away. The receiver
will then receive the information and it will perform the task according to the information.
(Refer Slide Time: 25:32)
Here the information is encoded in calcium. This process involves signaling initiation and
propagation. It propagates the calcium waves and the receiver performs the task according to the
calcium concentration.
(Refer Slide Time: 25:47)

By using this principle, we can deliver the drugs to the targeted cancer cells very specifically.

(Refer Slide Time: 26:01)

But still lots of challenges needs to be addressed because we want the system to be autonomous.
We do not want any human control. We want the system to be closed, that means no energy
supply from outside should be required. We also want the recycling of the carrier in molecules.
Moreover, we want slow and sustained delivery of drug molecules.

So as a summary of this lecture we have learnt what is DNA based nano machine and protein
based nano machine and we have also learnt nano scale communication. A lot of research is
going on in this field to explore the potential application of nano machines in various fields.

So I will end my lecture here. I thank you all for listening this lecture. I will see you in another
interesting lecture.

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