If you can recall back to my first post on influenza viruses (Spanish Influenza, Part 1), I said that viruses use stealth to obtain entrance to our cells. A virus’s ability to get inside relies on the virus’s ability to bind the cell. Proper binding, much like sliding a key inside a keyhole, unlocks the front door of our cell and ushers the virus inside. Influenza viruses rely on their coat protein hemagglutinin (key) to bind a specific molecule (lock) that hangs off the outside of our cell. The virus is looking for a sialic acid that can be presented one of two ways: α2,3 or α2,6. You don’t need to understand exactly what that means, but you do need to understand that α2,3 is found in birds, while α2,6 is found in humans.
Part 2 of our Spanish Influenza special explained where scientists were able to find RNA for the 1918 hemagglutinin protein. Through a series of steps (that aren’t important for this post), scientists were able to translate the RNA into amino acids, fold up the protein properly, and look at it. They studied the 1918 hemagglutinin from all angles and compared it to hemagglutinins from other influenza viruses.
From the post Protein Folding, I discussed the four levels that scientists use to describe protein structure. The first movie (movie! - those Ph.D. skills are paying off!) below shows you one hemagglutinin molecule from the 1918 virus (Movie 9.1). Secondary structure (alpha helices and beta sheets) can be easily seen. Also, this movie is showing you the tertiary structure of the protein. It answers questions like “How does it fold up in space?”, “Where are all those alpha helices and beta sheets?”, and “What about random regions with no secondary structure – where are they?” The movie offers you the ability to see the protein from several angles.
The fourth level of folding tells us how many individual molecules must come together to make a fully functional protein. In the case of hemagglutinin, the answer is three. Movie 9.2 shows you how three hemagglutinin molecules come together. This is what the protein looks like when embedded in the coat of the influenza virus and is fully capable of binding a soon-to-be infected cell.
Let’s take a moment talk about what the trimer looks like. The entire protein complex resembles a bouquet. The long stem region (which runs ~ 130 Å) is highlighted in Figure 9.1. Look carefully and you will see that each hemagglutinin molecule offers one long alpha helix to the stem and these three alpha helices all wind around each other forming what structural biologists call a coiled coil. This is a common way for alpha helices to come together in protein structures. The “flower” part of our bouquet is called the globular head region. The stem region and the globular head are known as protein domains, which are parts of the protein that fold up to form a structure independent of the rest of the protein. (I’ll go into more detail about domains in a different post.)
An area of great interest is the place where sialic acid binds. This falls within the globular head domain (Figure 9.1). Before we look at it, I want to highlight two key points:
1.
1. A sialic acid linked to a galactose in an α2,3 way looks significantly different than a sialic acid linked in an α2,6 way.
1. A sialic acid linked to a galactose in an α2,3 way looks significantly different than a sialic acid linked in an α2,6 way.
2 2. Humans have α2,6 linkages in their respiratory tract; birds have α2,3 linkages in their digestive tract. In order for an avian virus to infect a human cell, it must gain the ability to bind α2,6 linkages. This should correspond to a visible change in the protein at the site of sialic acid binding.
Okay – on to the area!! What does it look like? How does it compare to known avian hemagglutinins?
Figure 9.2 is directly from the paper and depicts the binding pockets for sialic acid. The deeper brown represents atoms set further back from the atoms in lighter brown. This is trying to give you a sense of depth to the pocket. The top panel shows 1918 hemagglutinin while the bottom panel shows an avian hemagglutinin. They look quite similar, yes. But remember that the bottom hemagglutinin can only bind α2,3 linkages but the top one has gained the ability to bind α2,6. Subtle differences must exist.
Check out the middle panel. That is a hemagglutinin from swine that can bind α2,6 linkages. The pocket is wide at the center. Compare again the top and bottom hemagglutinin. The top one is slightly wider than the bottom one. This widening of the pocket must be responsible for its ability to bind α2,6 linkages. It wouldn’t take much to widen this pocket – a simple amino acid change or two (which happens naturally) can widen it just enough to adapt the virus for human cell infection.
Of course, the widening of the pocket does not completely answer why 1918 influenza was so virulent. Our immune system “remembers” different proteins from invaders and hemagglutinin is one of them. However, look how little the pocket widened from an avian hemagglutinin and how dissimilar it looks to the swine hemagglutinin, which is fully adapted to binding α2,6 linkages. This protein had barely changed from a bird virus and it is easy to conclude that humans had never seen anything like it before.
From year to year, hemagglutinins that can bind α2,6 linkages vary slightly or are placed with new neuraminidases (Spanish Influenza, Part 2) to make new viruses. This is why we need a new flu vaccine each year – the viruses do change slightly. However, our immune system has seen similar proteins and can still wage a small scale attack on these viruses. In the case of 1918, this protein had never been seen before. Our immune systems basically looked at it and said “What the hell is that?” while the virus promptly decimated our cells.
Scientists continue to study this virus to obtain clues to why it was so virulent. Widening of the pocket, never seen before hemagglutinin, different cleavage loops (something I didn’t talk about), etc. are all clues to what create an especially deadly virus. If we know what the dangers are, then we can identify them before they become a widespread pandemic. At least, that is the theory!
Coiled coil: common structural motif in proteins where alpha helices wind around each other much like individual strands of rope wind around each other to make a stronger rope.
Domain: part of a protein that folds into its own structure and has its own function even in the absence of the remaining protein/amino acid sequence
References
Stevens et al. “Structure of the Uncleaved Human H1 Hemagglutinin from Extinct Influenza Virus.” (2004) Science 303, pgs 1866 – 1870.
PDB Code for 1918 hemagglutinin: 1RD8 (www.pdb.org)
Personal movies and figures were rendered in PyMOL.
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