Doing research in a protein lab, the most common question I get asked is ‘Are you doing it for the gains?’ (Gains is a colloquial term for building muscle through going to the gym and often by consuming large amounts of protein). If you’re like most people, on a day-to-day basis you may only really think about proteins as you cut into a steak or prepare a post-gym recovery shake. However, proteins do much more than help our biceps look good – they’re a family of compounds that are incredibly diverse and are essential for any living organism. Their importance is underscored by the Greek root of their name – protos, proteios – which means first, or primary. Proteins really do deserve to be in first place; they do everything from acting as messengers in our bodies (hormonal function), to catalysing reactions that make it possible for us to breathe and eat (enzymatic function), to supporting essential structures like skin and hair (structural function).
Perhaps the most interesting thing about proteins is that they are all made from the same building blocks: a set of twenty amino acids. From a scientific point of view this is exciting because it means there is a defined sequence that can be manipulated to affect the protein function. You may be thinking – only twenty amino acids? Easy! Just change every amino acid position of a protein until we get optimal sequences. But, consider a protein made up of only 6 amino acids (a VERY small protein). If we were to randomise completely at every position the total number of different proteins we would have to make and test is 64 million! (206). This means that part of our job as protein engineers is to either invent ways to quickly and cheaply test hundreds and thousands of proteins or to use rational design to guide our amino acid experimentation. However, using rational design is often easier said than done. For example, one of the proteins that my colleagues and I work with is a viral protein of 131 amino acids called CP3. When this protein is expressed in our lab it interacts with itself to form hollow spheres made up of 180 single proteins. As an engineer this assembly complicates my task because it means I must try to predict how changing one or more amino acids will affect the shape and assembly of the protein in 3D space – not an easy task.
Although there’s a lot we still don’t understand about the relationship between the structure and function of a protein, playing around with amino acids isn’t a complete shot in the dark. From the chemical structure of the 20 amino acids we can make educated guesses about how they will interact and behave. For example, only five of the amino acids are electrically charged so if you are looking to effect a protein’s reactivity, manipulating these amino acids may be a good place to start. Amino acids also vary in size and in hydrophilicity (how much a chemical “likes” to be around water molecules). Thus, depending on what your intended aim is – whether you want to change your protein’s reactivity, size, shape, 3D orientation, stability etc. – your amino acid selection strategy will vary. Looking again at the example of the CP3 protein, we can see from its sequence that it contains the interesting amino acid cysteine (Cys). When two cysteines come together they form a covalent bond which contributes to the stability of the large 180 protein complexes we observe. To increase the stability of CP3 one of our possible strategies includes adding more cysteines into the protein sequence to encourage more bonding between protein subunits. The beauty of creating symmetrical and repetitive complexes is that by simply adding one extra cysteine in my CP3 sequence the complex of 180 CP3s would potentially have 90 more bonding interactions!
How much does one amino acid really matter, you may ask? Sometimes, one amino acid can change everything. A perfect example is the MS2 virus protein CP (related to my protein of interest CP3) which also forms particles of 180 protein subunits. In the MS2 CP protein sequence of 140 amino acids, researchers recently found that one amino acid change at position 37 (from a Serine to a Proline) was enough to cause a complete switch in the geometry of the assembled protein particle and caused a size decrease of more than 30%. The real kicker? It wasn’t an amino acid that anyone would have predicted had any importance to the protein structure – and even now the reason it had such a big impact is anyone’s guess.
In the past few decades, the field of biochemistry and protein engineering has made huge strides, but there are many things we still don’t understand. While we can engineer proteins and we can literally build the “building blocks” of all organic life, a non-trivial proportion of this type of work still relies on luck. This is because we still don’t have the knowledge that allows us to accurately predict the function and role of every amino acid in every unique sequence. But, unlike engineers who build bridges and shopping malls and massive skyscrapers, scientists in the lab can tolerate a bit of chance and unpredictability. Smaller scale means less risk! Engineering proteins on the nanometre scale means working with particles that are 10-9 or 1,000,000,000 times smaller than the muscles forming your bicep. But don’t let the size fool you, the effect of the work that protein engineers do has the potential, quite literally, to change the foundation of organic life! But maybe we’ll just settle for inventing a way to turn proteins into “gains” more easily.
 Asensio MA, Morella NM, Jakobson CM, Hartman EC, Glasgow JE, Sankaran B, et al. A Selection for Assembly Reveals That a Single Amino Acid Mutant of the Bacteriophage MS2 Coat Protein Forms a Smaller Virus-like Particle. Nano Lett. 2016 Sep 14;16(9):5944–50.