Identification and Towards Verification of Riboswitch Candidates in the Human Gut Microbiome
Open Access
- Author:
- Mouck, Matthew M
- Area of Honors:
- Biochemistry and Molecular Biology
- Degree:
- Bachelor of Science
- Document Type:
- Thesis
- Thesis Supervisors:
- Philip C Bevilacqua, Thesis Supervisor
Joseph C. Reese, Thesis Honors Advisor - Keywords:
- RNA
Gut Microbiome
Riboswitch - Abstract:
- RNA is a highly diverse molecule able to perform a variety of functions, including gene regulation. Typically located in the upstream sequence of mRNA, riboswitches are able to respond to changing ligand concentrations in their environment by binding to a specific ligand and inducing a conformational change in the expression platform. This structural change is able to regulate transcription or translation, as different structures may form terminator or antiterminator hairpins as well as sequester or free the Shine-Dalgarno sequence. I hypothesize bacteria in the human gut may have evolved similar adaptive strategies via a riboswitch to respond to a human metabolite. Novel riboswitch classes have conventionally been found through computational techniques that exploit the high level of conservation of secondary structures. Therefore, I adapted a previously established computational pipeline to search for novel riboswitches in the gut microbiome for experimental validation.1 The pipeline requires sufficiently annotated genomes of gut microbiota and sorts proteins into groups based on their conserved domain. The upstream mRNA sequences of the proteins are obtained and grouped in accordance with their respective proteins. Consensus secondary structures are determined with a covariance model, and homologs of these structures are found in the original genomes and added to the matching group. Final consensus structures are produced and filtered to three candidates for validation by examining structural biology and functional biology. The three candidates are analyzed for changes in secondary structure via chemical probing with 2-methylnicotinic acid imidazolide (NAI). Due to restricted experimental timing, I cannot report conclusive results from these experiments. In future studies, these experiments should be finalized, and different candidates selected if none were indicative of riboswitch mechanistic activity.