Fun With Biology: Exploring The Molecule Of Life
“DNA is like a computer program but far far more advanced than any software ever created.” — Bill Gates, business magnate, software developer, investor, and philanthropist
Everything has the capacity to self-assemble. Any system, whether it is a living cell, a cell phone, buildings, planets, stars, or galaxies in the universe — they assemble based on the components, energy, and environment, and finally, get to the state of equilibrium.
Understanding Life Better
Talking about Biology, the most important components from which life is made are biological macromolecules, such as the DNA, RNA, proteins, sugars, and lipids, to name a few. Out of this list, nucleic acids (DNA, RNA) and proteins are the hottest topics of research because their structures encode the information that is critical for the continuity of life. The main purpose of fields like Biotechnology and Biological Engineering is to screen and explore the nature of such biomolecules and try to adapt and repurpose some of their purposes in the quest of a new function not yet found in nature!
The latest example of understanding this better is the COVID-19 pandemic. At this moment, the world is not immune to the novel coronavirus — so, can we come up with developing some new property in the existing structures to adapt to it? The biggest challenge, in the present times, is to develop a vaccine for this illness and its conventional process takes a lot of time. In such a case, is it possible to somehow trap this virus and degrade it till we get a more permanent solution? It would be really cool if we can develop some kind of technology for this, utilizing the naturally occurring biomolecules.
Ideality vs. Reality
In the recent past, the news of designer babies being created in China was doing rounds. According to some scientists, genes of choice can be put in a baby when it is still an embryo, after which they are born with different eye colors or other such characters. Hence, the ideal question is can we specify the functions of biomolecules and build it from scratch? But, the reality is that mankind cannot do it. We are still far away to properly engineer nature!
This sparks the question — what we can really do? We can realize the custom functions that exist in the synthetic molecular complexes and integrate them into the higher-order systems, which we can make in labs. In this way, we can apply these systems in science, technology, and health.
DNA — The Boss
As mentioned earlier, both proteins and DNA are crucial. Then, why choose DNA over proteins? Proteins are thread-like structures made up of amino acids. These threads can be folded in a variety of 3-D shapes. Given that multiple such shapes are possible, we still do not know how to predict or design tertiary structures of proteins very well. Moreover, these 3-D structures further form complicated quaternary structures.
The DNA can also do something similar. However, it is much easier to predict and play with it as compared to proteins. Why? Because, DNA is a simple duplex — a double-helical structure (like entangled noodles!). It is made up of nucleotides (adenine, thymine, guanine, and cytosine), which can very specifically self-assemble with each other using the Watson-Crick base-pairing. This base-pairing acts like a molecular glue and there are only two rules of complementarity — adenine pairs with thymine and guanine with cytosine. Based on it, we can explore unlimited structures arising from DNA.
DNA, short for Deoxyribonucleic acid, has three properties. The first is structural simplicity — it can be easily synthesized at a low cost. Then comes robustness and the last one is designability. There is software available to easily design and program different DNA structures. With enough complementarity, the DNA can progress from its basic form to assemble into beautiful 1-D nanowires/tubes and 2-D sheets/arrays at the nanoscale level. The next level is that of the 3-D nanocages/polyhedra. These can be visualized using (atomic force or electron) microscopes. The field which utilizes DNA as a building block is known as structural DNA nanotechnology.
The Power of Retrosynthesis
If a chemist is asked to synthesize a molecule, they generally break it into small parts that are easy to make, then synthesize and assemble them to generate the final molecule. This process is known as retrosynthesis. Now, a typical virus is icosahedral in geometry. Retrosynthesis can really help us if we want DNA to take this shape.
We can start by cutting the icosahedron into half. We get two cup-shaped structures, which are each formed from a single central vertex (with five arms). Using its DNA-forms and more such vertices along with the rules of complementarity as needed, we can get a geometry that can lead to exact the cup-shaped structures, which can then be assembled into the shape of the virus. Voila! The correct bioengineering approach can give us a DNA mimic of the virus.
This magic of DNA can lead to diverse architectures, viz., platonic solids, and Archimedian solids, in the long list of many others.
DNA Nanodevices: Origami with DNA
Many of us have seen our grandparents knit strands of wool into patterned sweaters, caps, mufflers, and so on. We can do the same with DNA as well! Developed in 2005, this technology is called DNA origami.
Basically, it starts with a naturally occurring long single strand of DNA, isolated from viruses or bacteria. Then some small strands, known as the staple strands, are added to the solution. These small pieces have the capacity to cause the long DNA strand to fold into a large variety of confined structures. All this happens inside a test tube, in a small machine called the PCR (Polymerase Chain Reaction). The 2-D origami shapes can further be folded into several 3-D designs.
The greatest advantage of this procedure is that these sequences of DNA encode important information and we are translating it into useful structures and devices, ranging from as few as five nanometers all the way to 100s of nanometers.
Much More Than Just Fun
The most basic question related to these structures is — are these just fancy or do they have some useful applications? Well, they have many uses, which can be broadly classified into three categories, the biosensors, bioimagers, and therapeutic agents.
A DNA-switch as a pH sensor is a first-generation biosensor developed by Yamuna Krishnan. It consists of a linear DNA that exists in an open conformation at a neutral pH (equal to seven) and we get a green signal (because fluorophores are present at the ends of this DNA). On the addition of a drop of acid, the structure changes to form a particular shape known as the i-motif structure, giving us a red signal. Let’s consider an example of a small organism called C. elegans, a 1mm long transparent nematode. After injecting these DNA devices inside it, we can see where they go. They are targeted to some particular cells since they are attracted by the receptors on these cells. Observations depict that this area is the liver of the organism. Apart from this, these nanodevices can be modified (by adding tags) in a way so that they travel to a specific region/site inside the body. This helps in the pH imaging of those cells at different time points, which can then be correlated with several diseases. This application is not just limited to sensing the pH, but it also maps different ions and metabolites in the cells.
The next interesting thing is the DNA mimic of the virus, which we discussed briefly some time ago. As mentioned earlier, the upper and lower cups, when mixed in a 1:1 ratio, form a cage. During this final step, an external agent like gold or magnetic nanoparticles, fluorescent polymers or proteins, drugs or antibodies, can be entrapped inside this cage. It can act as a very specific cargo delivery and bioimaging agent. Again, considering the case of C. elegans, a simple drug on injection inside the nematode gets diffused throughout its body. The same drug, when encapsulated inside the DNA cage, after injection gets targeted only to those cells that express receptors for this drug.
Imagine the situation of cancer. When we give chemotherapy to the patients, there is a bulk of medication that diffuses to other body locations. It is this phenomenon that causes damages such as eyebrow and hair thinning, skin and kidney shrinking, etc. Now, imagine a scenario where we can put these cancer-targeting drugs into cages made up of DNA and tag them such that they can reach specifically to the diseased cells, releasing the medication in the affected area. It will help in decreasing the side effects of chemotherapy drastically!
For this drug-DNA cage complex to reach and act on a specific site in the body, we need to give it the address of that location. It is known as tagging and is the process of coupling proteins, sugars, small molecules, peptides, and such, on the surfaces of the cage at a very specific place. Using computational modeling, we can develop atomistic models of these cages, which help us in understanding their precise geometry so that we can add the tags at desired locations. Through advanced microscopy, we can map various events inside the body, say, the journey of fluorophore tagged drug cages at the single-particle resolution, the size of the tumor growth, in the long list of many others. Earlier, this had been one of the major challenges in Biology — how to track the molecules going from outside to inside of the cells? DNA nanotechnology, which studies various biomolecule dynamics at a very high resolution inside the cell, is answering some of these questions now.
The Latest Application: DNA Structures Against COVID-19
In present times, the novel coronavirus disease is causing global destruction at an alarming pace. One of the biggest challenges in front of scientists is finding medications and developing an effective vaccine fast. Vaccines can be broadly classified into two categories — first is the small molecules, and the second is antibodies. The problem is that developing a treatment takes a lot of time since several tedious trials are involved. So, the question arises that can we do something during these times till a more permanent treatment option is accessible? Can we come up with some applications of these DNA nanodevices that can combat SARS-CoV-2?
Around the world, multiple studies are going on where scientists are trying to explore DNA nanotechnology to block this virus and degrade it. It is based on the logic of a DNA cage with enzymes inside it. The cage can engulf the virus and the enzymes can destroy it. If used efficiently, this technique can remove a considerable number of virus particles floating in the bloodstream and lungs, thereby providing some therapeutic relief to the patients. Most of this work is being done in the renowned biophysicist Hendrik Dietz’s lab in Germany, with some complementary ideas to implement it in India soon.
DNA origami is inspired by how the virus evolves in nature. A virus has three proteins that can form a trimer, which in turn, can polymerize to form an icosahedron geometry. Dietz’s lab exactly replicated this self-assembly of the virus utilizing DNA and came up with different buckyball structures. They initially made two cups of the buckyball, which self-assembled to form the entire entity. These have cavities from which the virus particles can enter and get trapped within them. Now, why will the virus come and sit inside this cage? We know some of the surface proteins on SARS-CoV-2 and we can use antibodies that recognize these proteins to lure the virus. These antibodies are tagged with small DNA strands that can attach to the inside of the DNA cage based on the rules of complementarity. This environment is loved by the virus, many of which get attracted and trapped inside the cage! After this, these viruses are inhibited by enzymes like proteases or degraded by proteins like chaperons. According to the initial lab studies, an increase in the concentration of antibody-DNA cages is leading to about 80–90% virus blocking.
The next steps to be taken in the future include the know-how of in-vivo neutralization of the capacity of different targets and pursuing the degradation of trapped virus.
To summarize, the interesting field of structural DNA nanotechnology has evolved constantly over the last 25–30 years. It started with different designs and has eventually progressed to the functionalization of these designs (with practical implementations). At present, this field is working at the interface of several biology-related domains and is focusing on biomedical applications for the betterment of humankind…
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This article is based on one of the sessions (delivered by Dhiraj Bhatia, faculty in the Biological Engineering discipline) of the Virtual Seminar Series by IIT Gandhinagar. It is an online program started by the Institute in the wake of the current pandemic as a means to engage the people so that they can learn about a diversity of topics from the comfort of their homes, in an interesting manner. (The 1st article of this series can be found here. The 3rd article is available here.)
