As Vinothkumar Kutti Ragunath prepared to move from Cambridge, UK, to Bengaluru, his friends lost no time in briefing him about his home country.
The microscope would not arrive easily, he was warned, as a large number of clearances would be necessary for its shipment. It would remain in a corner unboxed after its arrival, waiting for a room to be ready for its installation. Then there would be a long list of things to be taken care of: water, air conditioning, electricity….
He thought it would take at least 10 months for the device to start working. India had been trying to get the cryo-electron microscope (cryo-EM) for about five years. The cryo-EM had, during this period, set the world of biology on fire, and every country with a good research base was trying to grab as many of them as possible.
Buying a cryo-EM was not a simple task. The microscope cost Rs 45 crore, and an additional Rs 20 crore had to be spent on special buildings and supporting equipment. There was so much demand for the machine around the world that manufacturers had orders to fulfill for several years.
More than anything else, the microscope required expertise to operate. This was Ragunath’s role. “Without him, the microscope would be like a doorstop,” says Satyajit Mayor, director of the Institute of Stem Cell Biology and Regenerative Medicine (InStem) in Bengaluru. When the InStem management reached out to Ragunath in Cambridge, he had no plans to move to India.
“I did not think India would get the microscope,” he recalls. He had been working for 11 years at the MRC Laboratory of Molecular Biology in Cambridge. With 11 Nobel prizes and many famous scientists, this laboratory was one of the nerve centres of modern biology. It was also the lab where some of the technologies for the cryo-EM originated.
Ragunath’s mentor there, Richard Henderson, had won the Nobel prize in chemistry last year for developing the cryo-EM. He shared it with Jacques Dubochet and Joachim Frank.
Cryo-EM changed the face of biology research within a few years. “It has already completely revolutionised structural biology,” says Venkatraman Ramakrishnan, deputy director of the MRC Laboratory of Molecular Biology and Nobel prize winner in 2009. “It will continue to make tremendous progress.” As a major research nation, it was important for India to get the microscope and Ragunath. Other major nations had plenty of cryo-EM: the US had more than 60, the UK about 15.
China has 11, and has supposedly ordered a large number of them. Help for InStem came from the MRC laboratory itself. Henderson and Ramakrishnan persuaded Ragunath to return to India. It was Ramakrishnan who had first introduced the microscope to InStem during a talk there, and then convinced the science leaders in the country that India badly needed to invest in the microscope.
“It needed a push and pull to bring Vinothkumar here,” says S Ramaswamy, senior professor at InStem and project coordinator of the bio cluster grant from the Department of Biotechnology (DBT) that funded the microscope. “Richard and Venki provided the push.”
Ragunath moved to India in July last year. The building and other infrastructure were finished when the microscope arrived. By September, it was set up and ready for use. “I was surprised at how quickly things worked,” says Ragunath.
The 1.5 tonne microscope now sits in a new building at the InStem campus, in a special basement room that is shielded from vibrations. In the next room is a facility to freeze biological samples in an instant. In another room nearby sits powerful computers, which will analyse a terabyte of data coming from the microscope every day. All of them were formally inaugurated last month as a national facility by scientist CNR Rao, who himself had been one of the motivations behind the move to bring the microscope to InStem.
Through the Lens, Precisely
Structural biology is a well-developed field in India, and a large number of biologists around the country use the older technique of crystallography to study the structure of proteins and other biomolecules. Proteins are the enzymes that digest your food, the antibodies that ward off bacteria and viruses, the messengers that carry important molecules from one part of the body to another, the skeleton that provides support to the cells…. Almost all functions in the body are regulated by proteins.
Biologists think that there are two million different forms of proteins in the body, and they do not know precisely how they work in a large number of cases. “Understanding the structure of a protein is the first step towards knowing its function,” says Ramaswamy.
Understanding protein function is an important step towards tackling complex diseases. Several Nobel prizes have been awarded recently for working out the structure of important proteins, which continue to be the target of many drugs. Ramakrishnan had won the Nobel prize in 2009 for working out the structure of the ribosome, the cell’s intricate machinery for protein synthesis. The bacterial ribosome, for example, is the target of some antibiotics. About 90% of current drugs target proteins that straddle cell membranes. About 50% of drugs target a set of proteins called G-coupled receptors.
Structural biologists are an important set of people. Crystallography images proteins by bouncing X-rays off them. For six decades, it was a standard method for figuring out the structure of proteins. It is a sophisticated technique that works well for proteins that can be coaxed to arrange themselves neatly into crystals.
It is easy to crystallise small proteins but the task becomes increasingly difficult as the protein size becomes large. Many of the larger proteins do not crystallise well, which is why their structure is not known. Cryo-EM changed this situation very quickly. “It has made all sorts of structures possible that would have been impossible to do by crystallography,” says Ramakrishnan. At the MRC Laboratory, Ragunath used the cryo-EM to study the protein ATP synthase.
Biologists love this molecule. It sits across the cell membrane, with its head on the inside and the tail on the outside, directing the synthesis of ATP (adenosine triphosphate), another important molecule for biology. But ATP synthase is not an ordinary protein. It is a tiny and complex machine with several parts: an electric motor, a chemical motor, a pump and an enzyme, all shrink-wrapped neatly into one big protein molecule. What it helps to make, the ATP, is used for energy storage in the body. ATP synthase is too big for crystallisation.
It was one of the first molecules that biologists attacked with their new gadget. The cryo-EM provided beautiful images of ATP synthase. Biologists now know how it works, and it has now become a target for antibacterial, anticancer and other drugs under development. “For the first time,” says Mayor of InStem, “we have an insight into how the ATP synthase works at the atomic level.”
Ragunath changed course after he moved to Bengaluru, moving on to other classes of proteins. He found collaborators very quickly within the Bengaluru bio cluster where InStem is located. Several scientists were waiting to use the microscope to solve protein structures. Ramaswamy, a structural biologist, is a key collaborator. The two scientists together would be a resource for biologists in and around the city, and in other parts of the country.
The Bengaluru bio cluster has two additional institutions: the National Centre for Biological Sciences (NCBS) and the Centre for Cellular and Molecular Platforms (C-Camp). C-Camp is also an incubator of biology startups, some of which need to solve protein structures regularly for their work. The first company to use the cryo-EM facility is Bugworks, which is developing a new generation of drugs against antibiotic-resistant bacteria.
Bugworks already has two drug candidates that aim to stop the bacteria from making copies of itself. They target the proteins responsible for unwinding the DNA in the bacteria, thereby not letting it duplicate itself. Drug companies like Bugworks need to know how a drug candidate binds to its target protein, and the cryo-EM will provide an image of the drug-protein complex easily.
“We use cryo-EM to optimise the next generation of drugs,” says Santanu Datta, chief scientific officer, Bugworks. “X-ray crystallography will provide only a static picture.” At the Indian Institute of Science (IISc), a few kilometres from the bio cluster, assistant professor Tanveer Hussain is preparing to use the microscope for his research on protein synthesis. Hussain had used the cryo-EM in Ramakrishnan’s lab at Cambridge, while working on the initial steps of protein synthesis. He will soon get a smaller cryo-EM at IISc, which will be used for screening samples to be taken to the larger one at InStem.
Scientists in other institutions are preparing to use it too. The Department of Biotechnology will fund a few smaller cryo-EMs at Pune, Faridabad, and IIT-Delhi. “The cryomicroscope should be seen as a symbol of India’s entry into microscopy,” says K Vijay-Raghavan, former secretary, DBT. India could amplify the benefits of the investment through technology development, especially in big data techniques. The microscope is evolving rapidly, and future versions will have deep reach while current versions will get cheaper.
The technology parts of the cryo-EM — the electron gun, the detector, computation and so on — improved gradually over the years but made a quantum jump around 2012-13. This improvement made scientists move to the field in droves in the last three years. Henderson, who played a key role in developing the cryo-EM, has a few ideas about the immediate future of the technology.
“We expect improvements of a factor of 20 in the information content of each image in two to three years,” says Henderson. This means that you can get contemporary images with onetwentieth the effort, or make the same effort and get images that are 20 times better.
This future excites scientists, and structural biologists using other methods are moving into the new field. So much so that companies that make the microscopes — the Bengaluru device was made by ThermoFisher — cannot make them fast enough. “It is a very exciting time,” says Henderson. India is joining the bandwagon a bit late, but not too late.
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