High Affinity Binding of N2-Modified Guanine Derivatives Significantly Disrupts the Ligand Binding Pocket of the Guanine Riboswitch

Abstract

Riboswitches are important model systems for the development of approaches to search for RNA-targeting therapeutics. A principal challenge in finding compounds that target riboswitches is that the effector ligand is typically almost completely encapsulated by the RNA, which severely limits the chemical space that can be explored. Efforts to find compounds that bind the guanine/adenine class of riboswitches with a high affinity have in part focused on purines modified at the C6 and C2 positions. These studies have revealed compounds that have low to sub-micromolar affinity and, in a few cases, have antimicrobial activity. To further understand how these compounds interact with the guanine riboswitch, we have performed an integrated structural and functional analysis of representative guanine derivatives with modifications at the C8, C6 and C2 positions. Our data indicate that while modifications of guanine at the C6 position are generally unfavorable, modifications at the C8 and C2 positions yield compounds that rival guanine with respect to binding affinity. Surprisingly, C2-modified guanines such as N2-acetylguanine completely disrupt a key Watson–Crick pairing interaction between the ligand and RNA. These compounds, which also modulate transcriptional termination as efficiently as guanine, open up a significant new chemical space of guanine modifications in the search for antimicrobial agents that target purine riboswitches.

A Riboswitch-Driven Era of New Antibacterials

“…Remarkably, N2-phenoxyacetyl guanine and N2-isobutyrylguanine modified on C2 position of guanine presented a similar binding affinity to guanine for the B. subtilis guanine riboswitch, resulting in transcription termination. However, the inhibitory activity of these compounds on the bacterial growth has yet to be determined [ 112 ].…”

Riboswitches as Drug Targets for Antibiotics

“…In fact, different riboswitches employ distinct strategies to achieve a high affinity and an appreciable degree of selectivity. Some riboswitches, like the lysine and purine riboswitches, possess binding pockets with perfect shape complementarity to cognate ligands and encapsulate them almost entirely and thus exclude binding of related compounds [ 35 , 36 ]. Often, hydrogen bonding with the ligand is an important means of achieving high specificity.…”

SILCS-RNA: Toward a Structure-Based Drug Design Approach for Targeting RNAs with Small Molecules

“…The binding site architectures of two of the RNAs, TPP riboswitch (Figure S3B) and HCV-IRES-IIa (Figure S3C), involve intercalation of the ligand. Based on available RNA–ligand databases − and other available literature, ,,,, we have curated a database of ligands targeting these RNAs. The Supporting Information file ct2c00381_si_002.xlsx provides the details of the small molecules, while ct2c00381_si_005.pdf provides their 2D structures.…”