The Power of Protons

By: Chauncey Chen

Figure 1. Dr. Qi Zhang

Expanding the alphabet by 50% would have innumerable consequences on language and communication. New research into RNA has essentially done this for our understanding of this molecule foundational to life as we know it. At the heart of this research is Dr. Qi Zhang, a structural and molecular biologist at the University of North Carolina at Chapel Hill (UNC). Originally trained as a physical chemist at the University of Michigan, Dr. Zhang has distinguished himself in the biology field by deeply investigating the structure of RNA molecules. In his 15 years at UNC, Dr. Zhang has been named as the inaugural Arrel Research and Teaching Distinguished Term Professor as a result of the accomplishments of his research group.

When asked about why he studies RNA, Dr. Zhang cites its centrality to so many aspects of life: “RNA plays roles in all different directions. They really play a central role in regulation, anything you can think of. So that’s what we got excited about.” RNA is found in every living thing, and nearly every biological process is related to RNA. For example, messenger RNA (mRNA) carries the code for every protein a cell produces. Notably, pathogens, including bacteria and many viruses, also interact with RNA, meaning understanding it will further push the frontiers of medicine. In pursuit of this goal, the Zhang lab conducts a wide range of research, connecting the chemistry of RNA to fields like oncology, immunology, and virology.

Like many things in the natural world, RNA should not be understood as static. Taking a single scan of an RNA is not enough to understand it, since that fails to capture changes over time. In Dr. Zhang’s words, “We are not photographers, we are filmmakers.”¹ We know now that RNA can be modified based on changes to the acidity of its environment. Typically, around 50% of RNA has a proton, or hydrogen ion, attached at a pH of four, meaning that at physiological pH, there should be minimal protonation. However, Dr. Zhang’s lab discovered that in certain microRNAs, which are small segments of RNA responsible for regulation, 50% protonation was reached at a pH about six, much closer to physiological pH. Moreover, over 80% of over four hundred RNAs were found to have protonation events at physiological pH.¹ This means that the number of nucleotides , or unique building blocks, in RNA has increased from four to six, since the two nucleotides which can be protonated have very different properties in their protonated and unprotonated states. In turn, this represents an exponential increase in the number of possible sequences, much like increasing the alphabet by 50% would exponentially increase the number of words which could be made. And just as more words would expand the range of ideas which could be expressed, new sequences mean a greater variety of possible RNA structures and functions. Therefore, measurements must be taken at different acidities, in order to see protonation occurring. What was found was that certain structures were responsible for maintaining the local protonation state of microRNAs, allowing protonation in conditions that otherwise would not be conducive to it.² This then dictates the formation of new bonds and structures.

Figure 2. Protonation Sites of the SARS-Cov-2 5’ UTR in Yellow.

The study of the SARS-Cov-2 virus also underscores Dr. Zhang’s belief in the importance of collaboration in science. In order to determine the structure, the Zhang lab worked with a group in Germany that used NMR, a technique that works by aligning the nuclei of atoms in a molecule and then measuring the signals released as they return to their natural states. From these signals, the structure of the molecule can be determined with a high level of accuracy.¹ However, NMR struggles with large molecules like an RNA sequence with hundreds of nucleotides, and therefore, the team focused on one small section of the near the start of the virus’s genome, the 5’ untranslated region. Although this region does not code for proteins, it nonetheless is highly important in regulation; for example, in SARS-Cov-2, it is a replication signal.³ Building off their collaborators’ work, the Zhang lab found in the case of the SARS-Cov-2 virus responsible for Covid-19 was that the 5’ untranslated region had multiple protonation sites, as seen in Figure 2, corroborating the findings of the NMR. The presence of the protonation sites throughout the 5’ untranslated region makes it very sensitive to pH changes, and thus a target for further study “With our technology, we show them the blueprint,” Dr. Zhang said.¹ Now, armed with the “blueprint”, the group collaborating with the Zhang lab is following up on the newly discovered protonation sites to better understand their structure with high-precision NMR. Since the 5’ untranslated region is key to the survival of the SARS-Cov-2 virus, disrupting its structure, which the study has shown to be possible,³ could be an effective treatment for Covid-19. Indeed, the ultimate goal is to take advantage of the protonation using small molecules to develop therapeutics which can disrupt the structure and function of the virus.

Until recently, the effect of protonation on the structure of RNA was poorly understood. Dr. Zhang cites this as one of the main challenges his lab has faced. In fact, it took five years of work to identify and confirm that protonation was, in fact, the cause of the structural phenomena observed. Now that the “common language” of protonated RNA is widely understood, the possibilities for RNA research have expanded greatly since the understanding of nearly every RNA-involved process could be influenced by this development. For example, the Zhang lab is currently researching the effect of RNA protonation on cancer growth. As our shared understanding of RNA deepens, the potential for RNAtargeting therapeutics will grow with it, expanding far beyond the laboratory walls into hospitals and clinics. As the field develops, the Zhang lab will surely be at its vanguard.

References:

1. Interview with Qi Zhang, Ph.D. 2/18/26
2. Baisden, J. T.; Boyer, J. A.; Zhao, B.; Hammond, S. M.; Zhang, Q. Visualizing a Protonated RNA State That Modulates MicroRNA-21 Maturation. Nature Chemical Biology 2020, 17 (1), 80–88.
3. Toews, S.; Wacker, A.; Faison, E. M.; Duchardt-Ferner, E.; Richter, C.; Mathieu, D.; Bottaro, S.; Zhang, Q.; Schwalbe, H. The 5′-Terminal Stem–Loop RNA Element of SARS-COV-2 Features Highly Dynamic Structural Elements That Are Sensitive to Differences in Cellular Ph. Nucleic Acids Research 2024, 52 (13), 7971–7986