{'fraction_non_alphanumeric': 0.020582329317269075, 'fraction_numerical': 0.0015060240963855422, 'mean_word_length': 5.470779220779221, 'pattern_counts': {'":': 0, '<': 0, '<?xml version=': 0, '>': 0, 'https://': 0, 'lorem ipsum': 0, 'www.': 0, 'xml': 0}, 'pii_count': 0, 'substrings_counts': 0, 'word_list_counts': {'cursed_substrings.json': 0, 'profanity_word_list.json': 0, 'sexual_word_list.json': 0, 'zh_pornsignals.json': 0}}
Self-cleavage of the glmS ribozyme core is controlled by a fragile folding element

Significance Riboswitches are structured mRNA elements that regulate gene expression in response to metabolite ligands. The bacterial glmS riboswitch regulates a protein with a key role in cell wall biosynthesis. This riboswitch follows an unusual mechanism, acting as a ligand-activated self-cleaving ribozyme. Understanding how this RNA folds may aid development of antibiotics targeting it. We used optical tweezers to investigate this ribozyme riboswitch at the single-molecule level. We found that folding and function are intertwined in this system, with a dynamically unstable folding transition controlling catalysis. Furthermore, binding of the ligand fails to stabilize the weakly favored folded state. Hence, the glmS ribozyme does not follow the conventional model for riboswitch switching, in which ligand binding stabilizes a particular switch conformation. Riboswitches modulate gene expression in response to small-molecule ligands. Switching is generally thought to occur via the stabilization of a specific RNA structure conferred by binding the cognate ligand. However, it is unclear whether any such stabilization occurs for riboswitches whose ligands also play functional roles, such as the glmS ribozyme riboswitch, which undergoes self-cleavage using its regulatory ligand, glucosamine 6-phosphate, as a catalytic cofactor. To address this question, it is necessary to determine both the conformational ensemble and its ligand dependence. We used optical tweezers to measure folding dynamics and cleavage rates for the core glmS ribozyme over a range of forces and ligand conditions. We found that the folding of a specific structural element, the P2.2 duplex, controls active-site formation and catalysis. However, the folded state is only weakly stable, regardless of cofactor concentration, supplying a clear exception to the ligand-based stabilization model of riboswitch function.