 So this far we've talked about passive transport, we talked about active transport, we've talked about signaling, which is kind of transporting information. There are a huge number of other proteins. We have proteins that are responsible for sensing pressure so that they're mechanical sensitive, they react to stress or so, and there's a relatively large part of proteins that are merely anchored in the cells, but they're not meaningless for that reason. You have likely seen this lately. This is a virus that has these small crown-like particles on it, an aura, in this case SARS-CoV-2, the virus responsible for COVID-19. If you magnify that part and have a look at at least a schematic look at this virus, what happens on this side of the virus is that this is an RNA virus, so you have RNA on the inside, the genetic material that will be injected in the cell. For this virus to work, it somehow has to get that genetic material into the cell, otherwise it won't spread, and that happens with these red so-called spike proteins. They're anchoring to the cell and somehow helping the virus fuse with the cellular membrane, and when they fused with the cellular membrane, it will be able to release its genetic information and using enzymes to, well, merge that in the cell's factory and create more virus proteins so that we build more copies of this. The way that happens, the red part here is central, and you've seen that red part. Again, the red one is the part that's blue up here, the spike. The spike protein in the case of SARS-CoV-2 for all viruses, you have some sort of receptor that would like to bind to the host cell, and the spike protein on SARS-CoV-2 binds to the ACE2 receptor on our cells. The ACE2 is something called angiotensin converting enzyme. It's a receptor very common in our heart, lung cells, in testines, and a few other places. What somehow happens is that the red spike proteins bind the blue ACE2 receptor, attach its cell to the host, and then manages to fuse the entire membrane. Let's see at least schematically how that works, because we don't know all the details yet. What will in general happen for viruses, for instance flu2, is that you have an early step where you first just attach the viral particle to the host cell. That means that you're now in spatial proximity, and that's important because this is going to be a reduction in entropy to stay here, and we need something to counter that. So the binding energy helps us stay close to the cell. That binding occasionally leads to a release of another part of the protein, and it might be that we're cleaving off a domain or something. That will release so-called fusion peptides, and these fusion peptides are essentially small drills from the virus point of view. These small drills will attach themselves to the host cell membrane, and you now have a protein part that goes from the viral membrane all the way into the host membranes, which is causing an even closer attachment of the particles. Now, if these two membranes are right next to each other, it's still better for each membrane, there is still some water here, so it's still better for the first membrane to be separate from the second membrane. But if we keep them really close to each other, under some conditions we might be able to get over this free energy barrier and get the hydrophobic tails in contact with each other. When those hydrophobic tails are in contact with each other, you can actually show that that's a transition state, and suddenly it's going to be easier for the molecule to go in what we call the hemifused state. I think that's what you have in the middle on the lower line here. So what's now happened is that we would start out this way. Let's draw the first membrane there, and the second membrane there. They're just very close to each other. So this would be B, and that would be A. Now, the first thing that would happen is that we would have A and maybe B, something like that, right? So they're very close to each other, but they still haven't fused. In the next step, what happens is that we have A and we have B, and one of their membranes has fused, kind of. So there is now one part here where we just have phospholipids that can move freely between the two layers. So this is not quite, they're still facing each other here, but there's one component that's hydrophobic between them. You could think of that as, you could all, maybe it's easier to think of it this way. So you have a very tight radius there. So there's one component sitting together, but the A and the B parts are really one factor. This is going to be stable for a while, because now we're having hydrophobic interior, and that we relax. And eventually, though, we have quite a lot of curvature energy here. It's bad to bend the lipids. So eventually, this part will grow, and as this part grows, you will get to the point where we have a larger and larger membrane, and eventually, you're going to end up with the case where both the membranes have fused. Ah, how should we draw that? Something like that. And then we have full contact between the inner part of the virus and the inner part of your cell. It's full fusion, and the viral material can enter the cell. This is something that we've mostly had to understand with indirect models, trying to guess what happens with the fusion peptides and everything. But this particular process, we can actually simulate. Let me show you that. So some 10 years ago, Peter Castle was supposed to stand for it, and here is Dawkel, and worked on exactly these processes. What happens during viral influenza infection, and in particular, how do membranes fuse? So what Peter did is that he didn't try to reproduce this entire system. That would have been too costly. But he created an ideal system where you had one vesicle, so that is a spherical double layer of lipids, radius roughly 50 nm, and a second vesicle here. But instead of having the actual proteins, he was tethering them together with a small chemical linker, 8 to 10 atoms. And what's going to happen now when I start this simulation is that you will first see the vesicles attaching to each other, simply because there are lots of charges in those head groups. It's quite advantageous for them to interact. Eventually, they will start pushing out the water to the side. And at some point, you're not going to be able to see it on this side, but there will be a case where one tail in the vesicle just happens to get in contact with the hydrophobic tail in another vesicle. That's going to be the transitional state, and when that has happened, they move relatively quickly to this hemifused state. And once they're in that state, a very short time later, they refuse completely. Here we go. So first, they find each other, they're pushing out the water, they're going to the hemifused state, and there we go to the fully fused state. And now the contents of the left vesicle has mixed with the contents of the right vesicle. The advantage of using this in computer simulation is that then you can use fairly simple energetics to understand what's happening. For instance, you can start many simulations like this, try to identify what is the probability if you're starting from a particular state of either fusing or not fusing. And then you can again draw up the entire free energy landscape, saying that you're starting from two vesicles, the free energy will go down a bit when they're close to each other, but they're going to stay around for quite a while. Then there will be a free energy barrier as we move to a transition state, the two vesicles fusing. We will go down to hemifused state, which is better than the initial ones. And at some point there will be a second free energy barrier when we move all the way to a fully fused state. So here we would have the hemifused state and the fully fused. So what's the point of doing simulations like this? Well, many I would say. First, we make sure that we understand the process from a molecular point of view. Second, we can start to investigate what are the components, for instance, causing vesicles to attach. What are the time critical steps here? In this case, it turned out that we had to squeeze the water out of the layer between the two vesicles. Third, you can study what is the likelihood that things will get stuck in a hemifused state. This turns out to be related to the charges on the head groups. So depending on the lipid composition, it will be easier or harder to fuse, which is going to be super important, not just for viruses. Third, how do things depend, say, on the length of the lipid tails? And fourth, how do those fusion peptides work? We still don't really know. One way or another, we think that they're perturbing, we're inserting a small protein like a drill, and we think that that is perturbing the structure of the lipids to increase the likelihood that the lipid tail will stick out. Because when that lipid tail sticks out, that's how we reach this transition state. But to tell the truth, a bit of this is still speculation and both Peter and several other teams are still working on this. So do join them if you're interested in doing a PhD.