Poster abstracts
Poster number 51 submitted by Opeyemi Fatunbi
NUCLEOBASE protonation in an RNA Thermometer
Opeyemi O. Fatunbi (Department of Chemistry & Biochemistry, Ohio University), Md. Ismail Hossain (Department of Chemistry & Biochemistry, Ohio University), Erin R. Murphy (Biomedical Sciences, Heritage College of Osteopathic Medicine, Ohio University), Jennifer V. Hines (Department of Chemistry & Biochemistry, Ohio University)
Abstract:
Infectious diseases have claimed more than 15 million out of the over 57 million annual global deaths,1 and the second leading cause of global death has been associated with bacteria.2 Many pathogens responsible for these infections have RNA as their genetic material. Many pathogenic RNAs have non-canonical base pairs, which may be protonated with perturbed pKa towards neutrality. The pH dependence and base pair protonation have been reported to influence various biological processes, such as miRNA processing and viral frameshifting.3 Bacteria such as Shigella dysenteriae, which causes Shigellosis and accounts for about 1.1 million annual deaths, have the Shigella heme uptake (shu) system, such as shuT.4 An RNA thermometer (RNAT) is located within the 5’- UTR of shuT. Structurally, the shuT RNAT has non-canonical base pairs (C●U and U●C) that may influence its structure and function. Hence, to fully understand the structural and functional relevance of the non-canonical base pairs of the shuT RNAT, it is pertinent to determine their possible pH dependence, pKa shift, and protonation state.
Herein, we employed UV spectroscopy and fluorescence spectroscopy to measure the pKa of the shuT RNAT. We observed a clear pH dependence and one ionization event in the shuT RNAT. In addition, we observed a pKa shift towards neutrality, and we were able to identify the non-canonical base pair that may be protonated, responsible for the pH dependence and upward pKa shift. These findings suggest that the protonation of one of the non-canonical base pairs may be pertinent in the structural stability, function, and interactions of the shuT RNAT.
Understanding the pH-dependent behavior of RNAs is vital in the drug discovery process that targets RNA molecules. pH environment in which RNA protonation occurs should be considered when designing small molecules capable of interacting with RNAs. This knowledge can guide the development of more effective RNA-targeted drugs, thereby advancing the field of RNA-based drug discovery.
References:
(1) Gliddon, H. D.; Herberg, J. A.; Levin, M.; Kaforou, M. Genome-Wide Host RNA Signatures of Infectious Diseases: Discovery and Clinical Translation. Immunology 2018, 153 (2), 171–178.
(2) Zhang, C.; Fu, X.; Liu, Y.; Zhao, H.; Wang, G. Burden of Infectious Diseases and Bacterial Antimicrobial Resistance in China: A Systematic Analysis for the Global Burden of Disease Study 2019. The Lancet Regional Health - Western Pacific 2024, 43, 100972.
(3) Wilcox, J. L.; Bevilacqua, P. C. A Simple Fluorescence Method for pK(a) Determination in RNA and DNA Reveals Highly Shifted pK(a)’s. J Am Chem Soc 2013, 135 (20), 7390–7393.
(4) Wei, Y.; Kouse, A. B.; Murphy, E. R. Transcriptional and Posttranscriptional Regulation of Shigella shuT in Response to Host-Associated Iron Availability and Temperature. Microbiologyopen 2017, 6 (3), e00442.
Keywords: Shigella dysenteriae, RNA thermometer, pH dependence