Industry Insights with Yuning Chen on Recombinant Proteins

This episode of Share Science features Yuning Chen, PhD, an R&D manager at Sino Biological. Sino Biological has an extensive library of recombinant virus proteins known as the ProVir collection, and in this interview, Yuning shares the importance and value of these high quality reagents.

What are recombinant virus proteins and how are they made?

Viruses are composed mainly of protein and genetic material, which is either DNA or RNA. Proteins are important for viral functions like replicating, interacting with the host, and achieving correct assemblies. In general, directly working with highly potent viruses is dangerous. One way to get around this issue is using the recombinant format of these protein components in various studies.

“I think the importance of these recombinant virus proteins is that they can … serve as a surrogate for the real virus so we can interrogate different aspects of a certain virus [and] better understand [its] biology, physiology, and functions.”

Recombinant virus proteins are made by a technique called recombinant expression, which has been around for almost five decades now, I think. It’s a relatively old technique, but it’s improving all the time. In this process, we take the gene fragment that encodes the protein of interest (in this case, a protein from a virus) and put it into a vector, normally in the format of a plasmid.

Plasmids are circular DNAs you can find in nature, mainly in some bacteria. We take this plasmid that contains the gene of interest and put it into a host cell. Basically, we use cells to help us mass manufacture these proteins. The host cells can be prokaryotes like Escherichia coli, which is one of the main workhorses in this industry. We also have eukaryotic systems such as yeast, just like what you use to make bread, as well as insect cells and even mammalian cells.

“We have all kinds of host cells, and depending on the characteristics of the virus protein, we pick one that will give us maybe the best results.”

We insert the plasmid containing the proteome (i.e., the gene of the protein of interest) into the host cell which has a protein manufacturing machinery that essentially mass produces this virus protein. At the end of the culturing cycle, we can harvest the cells or the culture supernatants and purify the virus protein with either the cell lysate or culture supernatant. This way, we can have an infinite supply of virus proteins to conduct all kinds of research.

Why do we need recombinant virus proteins and how are they used in infectious disease research?

Like I mentioned earlier, directly working with viruses can be dangerous sometimes, especially those categorized as biosafety level four pathogens such as Ebola or Marburg virus. Since viruses mainly use proteins to carry out their various biological functions, proteins are very important tools for us to understand certain aspects of a virus. We can obtain recombinant virus proteins in relatively large quantities. Since they’re not a whole virus, they’re not infectious, giving researchers a lot of freedom to investigate biophysical properties. For instance, people have heard a lot about SARS-CoV-2 spike proteins, which is the protein that interacts with host cells. Instead of working with a live virus, researchers can safely work with the spike protein.

“We can also make the spike protein of [different SARS-CoV-2] variants and … establish how they interact with host cells.”

With different versions of this spike protein from different variants, scientists can assess how these proteins bind to receptors like ACE2 and its binding strength. Essentially, these recombinant virus proteins can give us tools to understand the physiology and biology of the virus without actually working with the virus. Although recombinant virus proteins are only part of the virus, some of the key physiological traits of the virus can still be sufficiently determined.

How are recombinant virus proteins used to develop vaccines and antiviral therapeutics?

This is a big topic: recombinant virus proteins can be used directly as vaccines. Vaccines come in different shapes and formats; some of them use inactivated or deactivated viruses, some of them use messenger RNA (mRNA), and some are protein subunit vaccines which are derived from recombinant virus proteins. I think there are several companies in China that are developing second generation recombinant subunit vaccines based on the SARS-CoV-2 spike protein.

“Recombinant virus proteins can indeed be used directly as vaccines or … as tools to help vaccine development.”

In terms of antiviral therapeutics, viruses have different parts. Let’s use SAR-CoV-2 as an example again, since I think people are pretty familiar with it. The spike protein is the main gateway for viral host interactions and therapeutic antibodies can be developed directly against it. Therapeutic antibodies are one form of antiviral therapeutics.

Also, scientists can take two recombinant proteins, solve their structure, and see if small molecule inhibitors fit within the space of those proteins. Once these inhibitors are identified, small molecule antiviral drugs can be made as well. I think recombinant virus proteins are very critical tools for developing these antiviral therapeutics.

What do you think the future might hold for this amazing technology?

I think people are paying more and more attention to vaccines that can be used to prevent different variants of a disease. For instance, we update our seasonal flu vaccines on a yearly basis, but it would be nice to have one vaccine that can help protect us against different influenza strains. The same goes for SARS-CoV-2.

“These universal vaccines are relatively difficult to make because viruses mutate all the time and we never know what mutation they are going to present to us.”

Taking SARS-CoV-2 for instance, from the original strain all the way to the Delta variant, there was only a small number of mutations occurring in the spike protein. Then came the Omicron variant, which I think completely shattered people’s views of a virus’ mutation capacity. Because viruses are so volatile, it is difficult to develop universal vaccines or broad-spectrum antiviral therapeutics. For SARS-CoV-2, an antibody can have a good effect on one particular strain, but when mutations occur, the epitopes at which the antibody binds to the virus protein are partially or completely destroyed, making the antibody less effective or completely ineffective.

“It’s a challenging process, but I think at least in terms of the universal vaccine front, we [have] some encouraging news.”

I think just last year, the FDA started a clinical trial for a form of a universal flu vaccine which could be quite promising. I believe it’s derived from the H1N1 formula, into which they graft the hemagglutinin protein of different strains. With one injection, people could develop antibodies against the different influenza strains.

I would say this breakthrough is only the first step. I think in the future, there will be more and more different types or formats of universal vaccines available to the public or under development. The mRNA platform is relatively new and has been proven to be quite effective in terms of vaccine development, and fast as well. I think with this technology and also with the more conventional recombinant subunit vaccine approach, we might be able to have a format of a universal vaccine against influenza or coronaviruses in the future.

In terms of these broad-spectrum antiviral therapeutics, viruses may mutate a lot, but they do have some key regimens in their genome or proteome that cannot be replaced. For instance, their RNA polymerase or proteases are relatively stable in terms of their mutation rates. I think scientists are now developing antiviral therapeutics against these more conserved targets. In the future, there might be either small molecules or antiviral therapeutics available for a broader spectrum of applications.

What resources are available to procure recombinant virus proteins and what specialties are required for their production?

Because this recombinant protein expression technique has been around for a long time, I think there are a lot of research institutes or biotech companies like Sino Biological that are producing these recombinant proteins. Of course, I think the research institutes and universities are the core of any field of scientific study. Some of the research labs might also have some resources in terms of these recombinant virus proteins they would be happy to share.

I think those are the two major resources where we can obtain some of these recombinant virus proteins. There has been a lot of technical know-how, knowledge, and experience gathered over the past 50 years. For a company or a lab to be able to make these recombinant proteins, they will need both hardware and software because these cell cultures require shaker flasks or even fermenters or gigantic bioreactors.

“Because the technique has been around for four or five decades, there are indeed a lot of host systems to select from.”

Each system has to be treated with respect and some experience is needed to use each expression system well and to solve problems which, believe me, do occur. We have done our fair share of recombinant virus protein development using different systems, so I can guarantee that there will be problems. When hardware and software are combined correctly and effectively, we should be able to produce all kinds of recombinant virus proteins for everybody.

Can you give us a brief introduction to ProVir from Sino Biological?

We are quite proud of this ProVir platform. Our founder actually worked at Merck for a long time; he was a leading scientist on one of their vaccine development platforms. When he returned to China and founded Sino Biological, he wanted us to focus on or at least pay some attention to the infectious disease front.

We created a lot of recombinant virus proteins from different viruses and disease areas and put them all together in this ProVir collection. We have an extensive collection for SARS-CoV-2, from parts of the spike protein and different spike proteins from different variants. We also have an extensive collection of influenza proteins.

“Scientists can take these proteins and mix them up and try to make … universal vaccines.”

We have a very extensive pipeline for upper respiratory disease viruses, as well. I think a lot of these proteins have been formulated into a high-throughput chip format to screen for various respiratory diseases. There are maybe close to 2000 recombinant proteins in this ProVir collection. Also, for some of these proteins, we have developed highly specific antibodies for diagnostic applications. These antibodies can detect these proteins in various assay formats.

I think at the beginning of the SARS-CoV-2 pandemic, because of the similarities between the SARS-CoV-2 virus and the previous SARS virus, we had a leg up in developing diagnostic tools and vaccines. Essentially, there may be pieces of a little bit of everything. It’s a big collection, so if someone is interested in researching virology or using recombinant virus proteins or antibodies, I think this is the go-to place for procuring these reagents. We do also have quite stringent quality control measurements so that these proteins and antibodies are of top quality.

“We hope the material in these collections [can] help scientists all over the world fight off current infectious diseases, and also help prepare [for] ones that might come in the future.”

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