BEACON Researchers at Work: Colicin and Immunity Binding: A Love Story.

This week’s BEACON Researchers at Work post is by University of Washington graduate student Carrie Glenney.

Proteins are the workhorses of life and they play an essential role in just about every biological function, from DNA replication to the immune system. Just as the structure of proteins can drastically change how proteins work, the way in which proteins interact with each other determines if and how essential biological tasks are performed. Protein interactions are determined in large part by their binding affinity and specificity for one another. The binding specificity of a protein describes its “choosiness”: does the protein interact with one specific protein or many different proteins? Binding affinity then determines the strength of the bond between interacting proteins: when the affinity is high, the proteins will “hug” even tighter and for a longer period of time.

A major evolutionary problem involves understanding how high affinity and high specificity protein interactions can diversify when they are critical for cell survival. Small changes in one protein may impact this binding intimacy to the detriment of the organism, so we expect changes to be rare.

Antimicrobial production in Escherichia coli exemplifies such a high-stakes game of protein diversification. These E. coli strains produce colicin proteins, narrow-spectrum antimicrobials that can kill other bacterial cells by destroying DNA, RNA, or by poking holes in the cell wall. When a cell produces these toxic colicin proteins, they must simultaneously produce a neutralizing immunity protein to avoid self-poisoning. Once released from the cell, the colicin can kill cells that do not produce their own immunity to the colicin. Neutralization of the colicin protein is essential to cell survival; thus, each colicin protein has its own unique immunity protein that binds it with high affinity and specificity. In fact, colicin-immunity protein pairs have among the highest binding affinities of all proteins. Despite these high stakes, colicin-immunity protein binding pairs have undergone extensive diversification: but how?

One of the first hypotheses to explain the diversification of colicins invokes positive selection. First, changes in the immunity protein confer an advantageous broadened immunity function: the protein can still bind its own colicin, and it can also bind another functionally distinct colicin.  Since colicin-producing E. coli naturally reside in ecologies where polymorphic populations are present, the ability to bind other types of colicin proteins could confer an advantage. Subsequently, complimentary changes occur in the colicin, leading to a novel colicin-immunity protein pair and the inability of the ancestral immunity protein to bind the evolved colicin protein.

I am currently using a method called directed evolution to explore this hypothesis. Directed evolution is a method used to engineer proteins with desirable properties. First, a library of protein variants is created using genetic mutagenesis. Next, this library is screened for the trait of interest. Mutants with the desired phenotype can then be isolated and sequenced to see which mutations have occurred. Using this method, I am able to create a library of immunity genes with mutations and screen them for broadened immunity function. I am particularly interested in exploring changes in binding affinities of broadened immunity mutants and measuring how this has impacted the fitness of the organism. I then plan to use broadened immunity mutants to select for the evolution of novel colicin proteins.

Understanding how proteins evolve is a fundamental problem in evolutionary biology and it transcends disciplines. I am excited about the opportunity BEACON provides to facilitate new ideas and form interdisciplinary partnerships with other researchers who are exploring the same types of questions.

I came to love science in a round-about manner. In high school and college, I had very little interest in science and I struggled to pass my math classes. I was working in social work when I discovered my fascination with the natural world through hiking and camping in the Pacific Northwest.

I share BEACON’s passion for increasing diversity in the sciences and making all science, particularly evolutionary biology, accessible to the public. Most people have an inherent curiosity about life and how it works, even if they don’t immediately think of it as scientific curiosity. Based on my own experience, I believe that harnessing this innate inquisitiveness is the gateway to an understanding of and excitement for science. In collaboration with BEACON, I hope to contribute to expanding science outreach and hands-on science opportunities for the public and K-12 students that might not otherwise have these experiences.

Many thanks to the Kerr lab at the University of Washington for their support of me and this research.

For more information about Carrie’s work, please contact her at cglenney at u dot washington dot edu.

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