Abstract
Riboswitches are structured cis-regulators mainly found in the untranslated regions of messenger RNA. The aptamer domain of a riboswitch serves as a sensor for its ligand, the binding of which triggers conformational changes that regulate the behavior of its expression platform. As a model system for understanding riboswitch structures and functions, the add adenine riboswitch has been studied extensively. However, there is a need for further investigation of the conformational dynamics of the aptamer in light of the recent real-time crystallographic study at room temperature (RT) using an X-ray free electron laser (XFEL) and femtosecond X-ray crystallography (SFX). Herein, we investigate the conformational motions of the add adenine riboswitch aptamer domain, in the presence or absence of adenine, using nuclear magnetic resonance relaxation measurements and analysis of RT atomic displacement factors (B-factors). In the absence of ligand, the P1 duplex undergoes a fast exchange where the overall molecule exhibits a motion at kex ~ 319 s−1, based on imino signals. In the presence of ligand, the P1 duplex adopts a highly ordered conformation, with kex~ 83 s−1, similar to the global motion of the molecule, excluding the loops and binding pocket, at 84 s−1. The µs–ms motions in both the apo and bound states are consistent with RT B-factors. Reduced spatial atomic fluctuation, ~ 50%, in P1 upon ligand binding coincides with significantly attenuated temporal dynamic exchanges. The binding pocket is structured in the absence or presence of ligand, as evidenced by relatively low and similar RT B-factors. Therefore, despite the dramatic rearrangement of the binding pocket, those residues exhibit similar spatial thermal fluctuation before and after binding.
Highlights
Riboswitches are structured RNAs that selectively bind metabolites for controlled gene expression
A riboswitch consists of two domains: an “aptamer” and an “expression platform” (Mandal and Breaker 2004; Winkler and Breaker 2003)
Binding of a metabolite to the aptamer domain leads to an allosteric conformational change that is transmitted to the expression platform via a structurally regulated “switching” sequence (Batey 2012), enabling control over gene expression (Peselis et al 2015; Wickiser et al 2005)
Summary
Riboswitches are structured RNAs that selectively bind metabolites for controlled gene expression. A riboswitch consists of two domains: an “aptamer” and an “expression platform” (Mandal and Breaker 2004; Winkler and Breaker 2003). Binding of a metabolite to the aptamer domain leads to an allosteric conformational change that is transmitted to the expression platform via a structurally regulated “switching” sequence (Batey 2012), enabling control over gene expression (Peselis et al 2015; Wickiser et al 2005). The mode of gene regulation by riboswitches is diverse, the ligand recognition by their aptamer domains is relatively conserved across classes (Mandal and Breaker 2004; Winkler and Breaker 2003). The structural dynamics of the add (adenosine deaminase) adenine (A) riboswitch has been studied The structural dynamics of the add (adenosine deaminase) adenine (A) riboswitch has been studied (Warhaut et al. Vol.:(0123456789)
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