Conformational Heterogeneity of the SAM-I Riboswitch Transcriptional ON State: A Chaperone-like Role for S-adenosylmethionine

摘要

Riboswitches are promising targets for the design of novel antibiotics and engineering of portable genetic regulatory elements. There is evidence that variability in riboswitch properties allows tuning of expression for genes involved in different stages of biosynthetic pathways by mechanisms that are not currently understood. Here we explore the mechanism for tuning of SAM-I riboswitch folding. Most SAM-I riboswitches function at the transcriptional level by sensing the cognate ligand— S-adenosyl methionine (SAM). SAM-I riboswitches orchestrate the biosynthetic pathways of cysteine, methionine and SAM, etc. We use base pair probability predictions to examine the secondary structure folding landscape of several SAM-I riboswitch sequences. We predict different folding behaviors for different SAM-I riboswitch sequences. We identify several “decoy” base pairing interactions involving 5’ riboswitch residues that can compete with the formation of a P1 helix, a component of the ligand-bound “transcription OFF” state, in the absence of SAM. We hypothesize that blockage of these interactions through SAM contacts contributes to stabilization of the OFF state in the presence of ligand. We also probe folding patterns for a SAM-I riboswitch RNA using constructs with different 3’ truncation points experimentally. Folding was monitored through fluorescence, susceptibility to base-catalyzed cleavage, nuclear magnetic resonance and indirectly through SAM binding. We identify key decision windows at which SAM can affect the folding pathway toward the OFF state. The presence of decoy conformations and differential sensitivities to SAM at different transcript lengths are crucial for SAM-I riboswitches to modulate gene expression in the context of global cellular metabolism.