Evidence that Electrostatic Interactions Dictate the Ligand‐Induced Arrest of RNA Global Flexibility

Abstract

Keywords conformation analysis; drug design; NMR spectroscopy; RNA Arresting flexibility to prevent RNA from undergoing functionally important conformational transitions is an established strategy for developing RNA-targeting therapeutics. [1] To understand the role of RNA flexibility in adaptive recognition [2] is also important for the rational design of small molecules that bind their RNA target with high affinity and specificity. [3] Yet few studies have quantitatively examined how RNA-binding therapeutics affect the flexibility of their RNA targets. As a result, little is known about the RNA-ligand interactions that are important for arresting different types of RNA functional flexibility. Here, we provide direct evidence by using NMR residual dipolar couplings (RDCs) [4] that electrostatic interactions play a primary role in dictating the degree to which small molecules arrest global motions in RNA.

Probing Na+-Induced Changes in the HIV-1 TAR Conformational Dynamics Using NMR Residual Dipolar Couplings: New Insights into the Role of Counterions and Electrostatic Interactions in Adaptive Recognition

“…Aminoglycosides bind TAR with affinities that correlate with their total number of positive amines (34). Our NMR studies suggest that the changes in the TAR conformational dynamics induced by small molecules are also correlated to their total number of positive groups (35).…”

Characterizing the relative orientation and dynamics of RNA A-form helices using NMR residual dipolar couplings

“…5a) [90][91][92][93][94][95] . Figure 5b shows these TAR conformational transitions as visualized through application of the presented RDC protocol 24,87,96,97 . Interestingly, one finds that molecules that contribute a larger number of cationic groups tend to stabilize more linear and rigid TAR conformations (Fig.…”

Structural plasticity and Mg2+ binding properties of RNase P P4 from combined analysis of NMR residual dipolar couplings and motionally decoupled spin relaxation

“…Watson-Crick regions of the two domains were modeled using an idealized A-form helix geometry, whereas the UUCG loop in domain II was modeled using an X-ray structure (Ennifar et al 2000) as previously described for TAR (Al-Hashimi et al 2002a). The idealized A-form geometry has been validated for nonterminal Watson-Crick base pairs using RDCs in a number of different contexts (Mollova et al 2000;Sibille et al 2001;Al-Hashimi et al 2002b;Pitt et al 2004Pitt et al , 2005Hansen and Al-Hashimi 2006;Musselman et al 2006). Shown in Figure 3A is the correlation between measured and back-calculated RDCs (16 and 17 for domains I and II, respectively) when independently fitting each domain to an order tensor.…”

“…NMR spectra were processed using NMRPipe/NMRDraw (Delaglio et al 1995) or Felix (Accelrys, Inc. 2002), analyzed using NMRView (Johnson and Blevins 1994), and overlaid using SPARKY 3 (Goddard and Kneller 2004). The 1 H-13 C splittings were measured in the divalent ion-free P4 sample as the difference in the upfield and downfield components of the C dimension, which were measured in separate subspectra using 2D 13 C-1 H S 3 CT HSQC experiments (Meissner et al 1997;Meissner and Sorensen 1999;Pitt et al 2005). The two sets of RDCs computed from the differences in splittings measured in the presence and absence of phage (15 mg/mL) yielded an RMSD of 2.4 Hz (data not shown).…”