• Alex Cloherty

A shield of sugar

Today on twitter, I saw a tweet from Tom Caniels (@tomcaniels, he’s great at highlighting new research on viruses, go give him a follow!) about a new pre-print manuscript that presents a detailed "Structure of the hepatitis C virus E1E2 glycoprotein complex”, as it is entitled. I was excited to see this - and it occurred to me that this is rather seasonal. Because with Christmas around the corner, what better time to talk about sugar?

Yes, you heard correctly - sugar. Glycoproteins, which are central to this story, are proteins that are linked to sugar groups, or to use the scientific terminology for this sweet stuff, ‘glycans’ or ‘carbohydrates’. These protein-linked sugars are a huge deal in the study of viruses like Hepatitis C (HCV).

The next big thing in HCV research is the aim to develop a preventative vaccine. HCV is a nasty virus in that it establishes a chronic infection that is difficult to get rid of entirely. If left untreated, this virus will slowly but surely destroy your liver, the organ that acts as your body’s Britta filter: sucking out all the unwanted stuff, like bacteria or poisons, from your blood. The goal with current HCV antiviral treatment is "sustained virologic response”. This is a fancy phrase that simply means that the genome of the virus, which is made of RNA, can't be detected in the patient's body anymore. For most patients – over 90% – this means that the virus never rears its ugly head again and that symptoms are considerably improved, which is fantastic. But, these drugs are also expensive, and don’t work for every patient. Thus, a vaccine would be a great asset to our anti-HCV armoury.

In order to design a targeted vaccine, we must first understand our opponent, and its potential reactions to the attack that a vaccine would constitute. In the case of HCV, our viral opponent is a mastermind in regards to evading our immune systems.

For one, the term ‘HCV’ actually describes many lightly different ‘quasispecies’. This is due to very rapid evolution of the virus. This virus is able to evolve so quickly because it uses a specific machine for its own replication - and that machine is kind of bad at its job. You might think that this machine, which scientists call the HCV polymerase, should be making exact copies of the genetic material that codes for the virus. After all, you wouldn't want loads of mutations in your own DNA, right? However, it makes a lot of mistakes. Rather than an experienced typist, this polymerase functions somewhat more like myself at 8am without coffee. What you may not have expected is that, for the virus, these ‘mistakes’ are actually beneficial. Each mistake results in a mutation, and although some of those mutations will inevitably result in faulty viruses, some of them will be beneficial. Some of them will act as a disguise.

When your immune system first ‘sees' a virus, the image of that virus is imprinted into the immune system's memory for use in future encounters. It is like a very exact ‘Wanted’ poster - except instead of a face, this ‘Wanted' poster details the proteins and sugars that make up the virus. But, if the polymerase typist introduces ‘mistakes’, or mutations, into the genetic material of virus progeny, the population of viruses in a community, or in your body, can change and evolve into many slightly different quasispecies. The genetic material of viruses codes for a sequence of proteins that will together make up the structure of a virus, and if the genetic code changes, the protein structure of the virus will, too. This means your body now has to recognize not one ‘Wanted' poster, but many slightly different ‘Wanted' posters for many slightly different versions of the virus.

To make things even more complicated, each mutation has the potential to change not only the protein structure of the virus, but also the sugars that are attached to those proteins. And now we come back to the ‘glycans’.

The main focus on glycans in regards to HCV is on the sugars attached to the proteins displayed on the surface of the virus. These two ‘envelope proteins’, i.e. proteins that are attached to the fatty membrane that makes up the virus surface, look like spikes covering the whole outside of the virus. Each of those individual spikes is made up of one copy of the E1 protein, and another copy of E2, locked in a molecular embrace and coated with sugars. In fact, when scientists checked the composition of these E1E2 complexes, sugar made up a whole third of the total weight!

Why all the sweet stuff? Well, HCV is well known for using a so-called ‘glycan shield’ - a shield of sugars - to make sure that its proteins don’t make too much of an impression on our immune system. As this review manuscript put it in more technical terms, HCV uses a "glycan shield that reduces the immunogenicity of the envelope proteins and masks conserved neutralizing epitopes at their surface”. This shield of sugar acts as a sort of blurring filter on top of the E1E2 complexes that are displayed on the viral surface, which makes our immune sytem gloss over them. This is probably in part due to the fact that during HCV replication in human cells, the sugars are actually made by the human cells, so our immune system recognizes these sugars as ‘human', rather than 'virus’. As a result, these sugars can serve as a shield that masks key parts of the virus that antibodies might otherwise recognize.

On top of acting as a physical shield, glycans may also help viruses hide from our immune systems by helping to orchestrate ‘viral envelope breathing’. That’s right, viruses breathe too! However, in the case of viruses, ‘breathing’ refers to the ‘opening’ and ‘closing’ of the proteins on the virus surface - like the E1E2 spikes of HCV - with changes in environmental conditions, like temperature or acidity. If a protein is ‘open’, that means it is open to attack by antibodies - its sensitive, recognizable parts are exposed. A closed protein, on the other hand, is all rolled up like a hedgehog covering its belly - it is hiding its vulnerabilities from the immune system. A recent paper indicates that glycans may be involved in regulating the ‘breathing’ of HCV's E1E2 complex, although more research is needed to figure out exactly how this works.

Since this heavily sugared E1E2 complex is the only viral protein on the surface of the virus, it is a key focus in the design of vaccines. Scientists want to design a vaccine that will be able to train the immune system to recognize this breathing complex, and destroy the virus with it. But remember - HCV is a master of mutation. So how can we possibly design a vaccine that will be able to protect us against the abundance of HCV quasispecies? The key is to figure out which specific chunks of protein and sugar that make up the E1E2 complex are most precious to the virus. With any microbe - or mammal for that matter - certain bits will be so key to the survival of that organism that they will be very unlikely to mutate. For example, some specific sections of sugar and protein of the E1E2 complex have such key, and niche, roles for the virus, like grabbing on to proteins on the surface of human cells to help the virus enter, that they cannot change too much due to the risk of losing their function. Because of their importance to the ‘survival' of the virus, these proteins and sugars are the least likely to be different across quasispecies.

You can compare this to dog breeds. Although daschunds are wildly different from Weimaraners, they were both bred to have the function of hunting dogs. So, although their body sizes and colouring are very different, they both still have eyes to spot prey, ears to listen to their companions, and four legs to run on. These are features that are key to their success as hunting dogs. Likewise, successful viruses need to hold on to some key features that allow them to succesfully infect host cells, and reproduce within those cells. If the proteins and sugars that make up those features mutate too much, that particular virus won’t be successful anymore, and will not be able to replicate itself at such a high rate as its more successful brothers and sisters. As a result, those particular mutations will then ‘die out’ of the population of viruses.

Step 1 for scientists, then, is to find those absolutely critical locations on the E1E2 complex, that should be similar across quasispecies, and design a vaccine that would help our immune system to target them. Because these critical locations should be more or less identical across quasispecies, such a vaccine could be the one vaccine to rule them all. And, what do you know, scientists have already found some of those key regions!

Alreadly, some so-called ‘broadly neutralizing antibodies’, which are highly effective at targeting HCV by sticking to these key regions, have been identified. And once they’ve been found, step 2 is to take a real good close-up look at them. And this is where the pre-print paper that started us off comes in.

It’s proved pretty tricky to get a good close-up view of the E1E2 complex because of its flexibility and its heavy sugar coating. Put simply, these features of the E1E2 complex make it much more difficult for our best technology to take a good picture. However, the authors of this pre-print manuscript have done a pretty darn good job. They present a high-resolution (i.e. very detailed) image of the E1E2 complex, including an analysis of the specific sugars in its sugar coating, and together with broadly neutralizing antibodies stuck to the E1E2 complex.

The authors were able to identify the exact places where those broadly neutralizing antibodies bind with particular precision, and they also saw some new features of the E1E2 complex that are helpful for scientists to be aware of in designing a vaccine. If you hear ‘stucture-based drug design’ in the news about the design of new COVID-19 treatments, this is exactly the strategy that they are talking about. The idea is to use the latest microscopy and data processing technology to get really detailed images of these really tiny pieces of virus (or bacteria, or whatever else we are designing a drug against), and then design a drug to target them specificially.

You can imagine this like designing a key to fit a lock. It’s a lot easier if you know exactly what the lock looks like, and can design each slot on the key to match the tumblers in the lock. That is what structure-based design allows us to do - rather than designing thousands of keys and hoping that one fits the lock, which is how drug and vaccine design was approached in the past. In the case of HCV, the job is not done yet - there is still no preventative vaccine - but with each study that sheds more light on the structure of this sugar-coated virus, and how it sneakily evades our immune system, we gain the background knowledge needed to design smart vaccines.

Until next time, Sugar coat it!


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