Cystic fibrosis is a genetic disorder that results in a thick and sticky mucus buildup in the lungs and pancreas, causing breathing difficulties and inefficient nutrient absorption. The thick fluid is especially detrimental to the lungs, damaging tissue and encouraging infection.
It is tragically common. Over 1,000 new cases are reported each year worldwide, and although life expectancy continues to increase, most treatments consist of daily therapies that target the symptoms of cystic fibrosis rather than the underlying cause.
A genetic disorder, such as cystic fibrosis, is caused by changes in the DNA that encodes a specific protein. DNA is made up of four possible bases, cytosine (C), guanine (G), adenine (A), and thymine (T) connected to one another via very strong bonds in a long strand. Two identical but antiparallel strands of DNA are able to bind to one another, C to G and A to T, which forms the double helical structure of our DNA.
Miniature machines in the cells — your proteins — are able to read the sequence of these four bases, and translate specific sequences into long chains of amino acids that fold to make new proteins. Simply put, DNA is an instruction manual made up of four possible letters (C, G, A, and T), which encodes all of the intercellular machines — the proteins — that allow us to live.
It is estimated that all of the DNA in a single cell in your body is 3 meters long, tightly coiled again and again in an ordered array and crammed into the nucleus which is about 6 micrometers in diameter. That is around 3 billion nucleotides per cell. Shockingly, a genetic disorder such as cystic fibrosis can be caused by a single base pair change out of that 3 billion if it is unfortunate enough to occur in the right place.
In the case of cystic fibrosis, one of the most common mutations it is a loss of three base pairs in the region of DNA that encodes the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR for short). The CFTR is a protein that spans the membrane of cells in your epithelium. It is responsible for pumping chloride ions across the cells that make up your lungs, pancreas, digestive tract, and other mucus-covered tissues. The three-base pair loss in the cystic fibrosis mutation causes the deletion of the 508th amino acid of the CFTR.
When the CFTR gene and its mutation were identified in 1989, the question posed to researchers was what loss of the 508th amino acid of CFTR does to its structure. In general, changing even a single amino acid can completely change a protein’s shape, abolishing its function. Therefore, in theory, if you could see the shape of the mutated protein and compare it to the normal protein, you might be able to make a drug that can return the protein to the correct shape. But how do you see something that is smaller than a wavelength of light? We can answer this by looking at a thought experiment suggested by Alexander McPherson.
Suppose I asked for your help drawing a picture of my brand new Toyota Tundra, which is invisible. As a determined artist, you might overcome your initial shock and then feel out the surface of the truck, climbing over the top of it, feeling the wheels, windshield, and hood. From this spatial map, you could try to sketch a fairly reasonable representation of my vehicle.
But what if I told you that you could not touch my Tundra, and had to maintain a distance of at least five feet from it at all times? That would be a bit trickier. Perhaps you get clever and return with a bucket full of tennis balls, and begin walking around my vehicle, throwing each ball with a specific velocity and angle at the truck and noting the trajectory and speed the ball has when it returns. If you were to do this thousands of times from many different angles, you might be able to reconstruct a general representation of the truck’s surface. If you used something even smaller, say bouncy balls, you would be able to distinguish even finer details; maybe the door handles and windshield wipers.
Now what if I told you the real reason you cannot see my new Toyota Tundra is because it is incredibly small — as small as a protein in fact? To see the large Toyota Tundra with increased resolution, you needed to throw projectiles that were much smaller than the object you wanted to study. The same is true now. If you were to “throw” something smaller than the protein-sized Tundra, such as electrons or X-Rays, you will be able to note how they bounce back and determine the surficial distribution of the object. This is exactly what scientists do in the two most popular methods for determining the structure of macromolecules such as proteins and DNA: X-Ray Crystallography and Cryo-Electron Microscopy.
X-Ray Crystallography and Cryo-Electron Microscopy have allowed researchers to solve the structures of thousands of macromolecules, including the common mutant forms of CFTR that lead to cystic fibrosis. In fact, researchers were probably shocked to discover that the CFTR protein retains its structure at its functional site despite the deletion of amino acid 508. Instead, the amino acid deletion unfolded a functionally irrelevant section of the protein, causing cell machinery to retain and destroy the misfolded yet fully functional protein.
Armed with this information, pharmaceutical companies began developing drugs that can help refold the CFTR protein. More structures of CFTR mutants led to the development of the drug Ivacaftor, which was called “the most important drug of 2012” for its ability to treat the underlying cause of cystic fibrosis.
Considering the medical importance of protein structure, it seems fitting that the 2017 Nobel Prize in Chemistry was awarded to Jacques Dubochet, Joachim Frank, and Richard Henderson for the improvements they made to imaging biomolecules via cryo-EM. The order of events I’ve given for the CFTR mutation: gene identification, mutation location, protein structure, and drug target, is one that is becoming increasingly common in the field of structural biology, and it is all thanks to our ability to see the proteins that we need to target.