Biochem research with Meghann Kasal

 

This week, I talked to Meghann Kasal ’17, who does research with Professor Stephen Miller in the biochemistry department. Kasal’s work centers around bacterial communication, specifically with a process called quorum sensing. In quorum sensing, bacteria produce molecules called autoinducers. The greater the density of bacterial cells, the greater the concentration of autoinducers, and past a certain threshold, the autoinducers are detected by the bacteria and result in a change in gene expression. This process allows for rapid, community-wide communication, used for things like bioluminescence (bacteria that glow) and biofilms (bacteria that stick to each other and form a film, often a major culprit in chronic bacterial infecitons). This communication, explained Kasal, can occur both intra-species (within the same species) or inter-species (between different species). Kasal focuses on inter-species communication. Quorum sensing in bacteria has strong implications for antibiotics, among other important future directions, so figuring out the mechanism behind it is hugely useful.

Kasal applied to do research in the biochemistry department with Professor Miller last summer after taking biochemistry in the spring. Coming into the lab, Kasal’s research would focus on a family of inter-converting molecules called AI-2 that are used in quorum-sensing communication in bacteria. AI-2 consists of one binding ligand that can convert between two known forms, although Khasal believes the molecule can probably convert between more than just those forms. AI-2 is special, both because it is produced by and binds to many different types of bacteria. One receptor that binds AI-2 is called the LsrB protein. Many bacteria can synthesize AI-2 using an enzyme called LuxS, and bind AI-2 by the LsrB receptor. Because of this, the greater the concentration of bacteria that can produce AI-2, the greater the concentration of AI-2 is, which enables the quorum-sensing communication that Kasal studies.

Kasal’s work was to identify receptors on novel species that could potentially bind  AI-2, and to figure out if they could bind the molecule based on the structure of the receptors. Kasal found potential LsrB-like receptors on species that are likely to bind AI-2 by identifying the extent to which the binding sites on the novel receptors were conserved. The more amino acids at binding sites that are conserved between the known LsrB receptor and the putative receptors on the novel species, the more likely the receptor is to bind AI-2. They had some bacteria species that had many binding sites conserved, and some with fewer. One species, said Kasal, only has 3 of the 6 key binding sites conserved on the receptor protein. However, Kasal said they still have hopes that it will bind, because the changes in amino acid sequence are modest compared to the known receptor proteins. Additionally, AI-2 itself is a molecule that is known to be able to adapt to its receptor, so if AI-2 can convert to other forms that have not yet been discovered, it may be able to bind many more species than previously thought.

Kasal went through a process of designing plasmids (small, independent bundles of the relevant genes for the receptor protein) and amplifying them using PCR, or polymerase chain reaction. PCR replicates a gene sequence of interest through a cycle of heating and cooling during which DNA polymerase, an enzyme used in DNA replication, creates a chain reaction of DNA replication at the desired site. The heating separates the two strands of the double helix, creating two template strands. When the temperature is lowered, the polymerase uses the strands as primers to replicate the sequence. The end result is thousands to millions of copies of the desired DNA sequence.

After doing PCR, Kasal inserted her genes into vectors (molecules that carry the protein) and transformed them into bacterial cells of different species to end up with new bacterial species that now contained the putative AI-2 receptors. In one species, she used two types: a LuxS+ strain, meaning it produced AI-2, and a LuxS- strain, meaning it did not produce AI-2.

After doing some trials, Kasal has found that three of her identified receptors have so far exhibited binding to AI-2. They have yet to do trials on the protein that only has 3 of the 6 binding sites that is conserved, but Kasal has hopes that it will work. The next step, she said, is figuring out the exact structure of each receptor protein and the type of AI-2 molecule binding to it. This can be done using a process called X-ray crystallography, in which the protein is grown into a crystal and then shot through with X-ray beams. The diffraction pattern — or the specific angles and orientations of the beams — can be converted into a three-dimensional model of the exact structure of the protein. Then, said Kasal, they can compare the structure of the protein to the predicted structure that their sequence gave them. Kasal also hopes that they can do this for a receptor protein that is bound to AI-2, because then they can observe the type of AI-2 molecule that is bound, and maybe even identify a new form of the molecule. The long-term implications for this are exciting: Kasal said that understanding the process by which bacteria can communicate in these large-scale ways has huge medical applications.

“If we can basically hijack bacterial communication, then we can control bacterial levels, and that can then be applied to, for example, the human gut microbiome, and it can be used alongside antibiotics and other treatments,” she said.

Kasal told me she knew she wanted to be a biochemistry major since early in high school. She loved chemistry in high school and especially loved organic chemistry here at Swarthmore, but has also loved biology. Biochemistry is, for her, the perfect match of those major interests.

“In my head, just combining [biology and chemistry] and looking at biochemistry is to me the way that I can put those interests together,” she explained. “I look at chemistry and I see all these ways to figure things out, but then I see bio and I’m like, I can use these realistically in these applications. I like that kind of detail, but I like studying it at the level of the organism.”

Khasal said she finds research specifically interesting because she likes to be able to solve a problem, “basically from step 0.” She likes not only the problem-solving aspect, but also the idea that she’s in the lab herself, carrying out the experiment that she designed.

Kasal expects to spend the next year and a half at Swarthmore working on the crystallization process for the AI-2 receptors, and writing her thesis on this research. After that, she plans to go to graduate school to pursue a Ph.D in a biochemistry-related field, and then she wants to become a professor. For Kasal, being able to pursue research while also being able to teach about her passion is the dream.

“I really like the idea of doing research and continuing research my whole life, but teaching is really fun for me and I really want other people to have that experience where they look at biochemistry and say, wow this is awesome,” she said.

 

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