The Second Class of Tetrahydrofolate (THF-II) Riboswitches Recognizes the Tetrahydrofolic Acid Ligand via Local Conformation Changes

Abstract

Riboswitches are regulatory noncoding RNAs found in bacteria, fungi and plants, that modulate gene expressions through structural changes in response to ligand binding. Understanding how ligands interact with riboswitches in solution can shed light on the molecular mechanisms of this ancient regulators. Previous studies showed that riboswitches undergo global conformation changes in response to ligand binding to relay information. Here, we report conformation switching models of the recently discovered tetrahydrofolic acid-responsive second class of tetrahydrofolate (THF-II) riboswitches in response to ligand binding. Using a combination of selective 2′-hydroxyl acylation, analyzed by primer extension (SHAPE) assay, 3D modeling and small-angle X-ray scattering (SAXS), we found that the ligand specifically recognizes and reshapes the THF-II riboswitch loop regions, but does not affect the stability of the P3 helix. Our results show that the THF-II riboswitch undergoes only local conformation changes in response to ligand binding, rearranging the Loop1-P3-Loop2 region and rotating Loop1 from a ~120° angle to a ~75° angle. This distinct conformation changes suggest a unique regulatory mechanism of the THF-II riboswitch, previously unseen in other riboswitches. Our findings may contribute to the fields of RNA sensors and drug design.

Bifidobacterium bifidum SAM-VI Riboswitch Conformation Change Requires Peripheral Helix Formation

“…After precipitation at −80 • C for 30 min, the RNA was pelleted by centrifugation at 12,000 rpm for 30 min at 4 • C. The pellet was recovered and resuspended in 10 µL of ddH 2 O. The SHAPE experiment was conducted following procedures described elsewhere [70][71][72]. Reverse transcription was carried out using a FAM-labeled DNA primer.…”

Riboswitches, from cognition to transformation

“…The THF-II riboswitch was firstly reported four years ago, and it mostly exists in Gram-negative Rhizobial bacteria while THF-I mainly exists in Gram-positive bacteria [ 38 ]. The THF-II riboswitch was demonstrated to undergo only local conformational changes after ligand binding, which provides a unique viewpoint on the regulatory mechanism of riboswitches [ 39 , 40 ].…”

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