Proteins have evolved to carry out a vast array of functions and, as we’ve discussed in previous posts on metagenomics, researchers can mine this natural diversity to find proteins that catalyze reactions useful for applied and therapeutic purposes. Indeed, researchers can find proteins that do everything from clean stains, to edit genomes, to detect DNA sequences.
Even with this vast natural diversity, it’s not always possible to find proteins with the precise function or optimal combination of characteristics that researchers want. While they may find very similar proteins, researchers often have specific purposes for which no currently known or naturally occurring proteins are ideally suited.
This is where protein engineering comes in. That is, researchers know enough about how proteins work and how they’re made to create new proteins with functions that are more specifically suited to their practical needs.
In this blog post, we’ll cover just a few of the many reasons that scientists engineer proteins. In a future blog post, we’ll discuss some of the techniques scientists use to achieve their protein engineering goals.
Changing substrate specificity
Proteins and, more specifically, enzymes catalyze many different kinds of chemical reactions – everything from joining amino acids together to breaking apart sugars for energy. For applied purposes, scientists often need proteins that catalyze a particular chemical reaction, say the reduction of an alcohol into a fuel-like alkane. While they may find natural enzymes that catalyze a similar reaction, they may not be able to act upon the substrates that scientists want them to.
For instance, scientists may be able to find many enzymes that catalyze the reduction of long alcohols into long alkanes. However, for the purposes of creating a biofuel, they might want an enzyme that catalyzes the reduction of shorter alcohols into alkanes. With an understanding of how natural enzymes interact with long alcohol substrates, they may be able to use protein engineering techniques to tweak the natural enzymes so that they act on shorter alcohol substrates.
In the case of Cas proteins, which catalyze the targeted nucleic acid-cutting reactions critical to CRISPR genome editing, scientists might have a particular DNA sequence that no known Cas protein is capable of editing with the required precision. These scientists can use their protein engineering know-how to alter the Cas protein so that it can edit the DNA sequence in the desired fashion. This would open up otherwise un-targetable regions of the genome to genome editing.
Enhancing reaction rates
In other cases, researchers may find a natural protein that is capable of catalyzing their desired reaction utilizing their desired substrate. However, the natural enzyme may only catalyze this reaction very slowly. In the example of biofuel production used above, if an enzyme can catalyze the reduction of a short alcohol into a short, fuel-like alkane, but can only do this once an hour, researchers might like an enzyme that works much faster. They can use protein engineering techniques to alter the structure of natural enzymes and potentially speed up the reaction.
Similarly, if a researcher finds a Cas enzyme that is capable of editing a desired DNA sequence but only very slowly, this enzyme would not be very useful in a clinical setting. For therapeutic purposes, it’s generally desirable for Cas enzymes to be present in cells for a short period of time and at low levels. Thus, if scientists use protein engineering techniques to speed up the Cas enzyme-mediated reaction without compromising specificity, this would make the Cas enzyme much more useful for CRISPR genome editing.
Protein design to alter thermostability
Enzymes often function most effectively at specific temperatures. Changing the temperature of a reaction just a few degrees can drastically alter how efficiently an enzyme catalyzes a reaction and changes in temperatures can sometimes even break enzymes.
It is sometimes possible to find organisms that produce enzymes that work at the temperatures researchers want. For instance, an enzyme that’s commonly used in the polymerase chain reaction (PCR) was isolated from a bacterium that grows in hot springs. This enzyme is active at the high temperatures required for the polymerase chain reaction.
Often, it is difficult to find enzymes that function at temperatures required by researchers. Here they can use protein engineering techniques to swap out amino acids in the protein to change how stable or functional it is at different temperatures.
For CRISPR genome editing, researchers generally want their Cas proteins to function best at body temperature. Thus, if researchers isolate Cas proteins that can target the DNA sequences they would like to edit, but these enzymes do not function well at body temperature, they can use protein engineering techniques to make these Cas proteins more functional in the body for more efficient genome editing.
For CRISPR diagnostics, the ability to function at a higher temperature is often preferable. This can lead to higher reaction rates and faster detection. Thus to optimize CRISPR diagnostics, researchers might want to engineer their proteins for stability at high temperatures.
Building on natural diversity with protein engineering
Even with the amazing diversity of proteins found in nature, researchers can enhance protein function for a variety of applied purposes using protein engineering. In a future blog post, we’ll discuss how researchers engineer proteins for new and altered functions.