Functional Roles of Chelated Magnesium Ions in RNA Folding and Function

Abstract

RNA regulates myriad cellular events such as transcription, translation, and splicing. To perform these essential functions, RNA often folds into complex tertiary structures in which its negatively charged ribose–phosphate backbone interacts with metal ions. Magnesium, the most abundant divalent metal ion in cells, neutralizes the backbone, thereby playing essential roles in RNA folding and function. This has been known for more than 50 years, and there are now thousands of in vitro studies, most of which have used ≥10 mM free Mg2+ ions to achieve optimal RNA folding and function. In the cell, however, concentrations of free Mg2+ ions are much lower, with most Mg2+ ions chelated by metabolites. In this Perspective, we curate data from a number of sources to provide extensive summaries of cellular concentrations of metabolites that bind Mg2+ and to estimate cellular concentrations of metabolite-chelated Mg2+ species, in the representative prokaryotic and eukaryotic systems Escherichia coli, Saccharomyces cerevisiae, and iBMK cells. Recent research from our lab and others has uncovered the fact that such weakly chelated Mg2+ ions can enhance RNA function, including its thermodynamic stability, chemical stability, and catalysis. We also discuss how metabolite-chelated Mg2+ complexes may have played roles in the origins of life. It is clear from this analysis that bound Mg2+ should not be simply considered non-RNA-interacting and that future RNA research, as well as protein research, could benefit from considering chelated magnesium.

Protein-Based Virtual Screening Tools Applied for RNA–Ligand Docking Identify New Binders of the preQ1-Riboswitch

“…Another aspect worth consideration is the functional role of ions. While a diffuse ion atmosphere masks the negative charge of the RNA backbone, especially free and metabolite-bound Mg 2+ ions play an important role in RNA folding, stabilization, and protection from degradation. , However, the identification of explicit (Mg 2+ ) ion binding sites in RNA is not trivial as only high-resolution X-ray structures allow for accurate structural determination. Furthermore, because of the high salt concentrations in crystallography, ion types and binding sites might differ from native conditions.…”

An in Vitro Cytomimetic of In‐Cell RNA Folding

“…However, typically RNA folding is studied in vitro, in dilute solutions, in the absence of other biomolecules; conditions that do not replicate the native cellular environment. Cells contain macromolecules such as proteins, nucleic acids and complex sugars with total concentrations of 50–400 g/L, resulting in steric “macromolecular crowding” and non‐steric “cellular sticking” interactions that are not reproduced by dilute in‐vitro solutions [9–15] …”

“…Several recent efforts to mimic the cellular matrix in vitro have been made by isolating “macromolecular crowding” or “cellular sticking” effects [6,12,16–21] . Artificial crowding agents, such as Ficoll and polyethylene glycol (PEG), are common ‘inert’ macromolecules used to simulate cellular crowding in vitro [19,22] .…”

Rational engineering enables co-crystallization and structural determination of the HIV-1 matrix-tRNA complex

“…If the sample shows heterogeneities, please refer to troubleshooting problem 3 below. Note: Mg 2+ is critical for RNA folding and stability ( Yamagami et al., 2021 ). To ensure that the concentration of Mg 2+ remains constant during the electrophoresis a peristaltic pump can be used to cycle and mix the running buffers.…”