Understanding the role of intermolecular interactions between lissoclimides and the eukaryotic ribosome

Abstract

Natural products that target the eukaryotic ribosome are promising therapeutics to treat a variety of cancers. It is therefore essential to determine their molecular mechanism of action to fully understand their mode of interaction with the target and to inform the development of new synthetic compounds with improved potency and reduced cytotoxicity. Toward this goal, we have previously established a short synthesis pathway that grants access to multiple congeners of the lissoclimide family. Here we present the X-ray co-crystal structure at 3.1 Å resolution of C45, a potent congener with two A-ring chlorine-bearing stereogenic centers with ‘unnatural’ configurations, with the yeast 80S ribosome, intermolecular interaction energies of the C45/ribosome complex, and single-molecule FRET data quantifying the impact of C45 on both human and yeast ribosomes. Together, these data provide new insights into the role of unusual non-covalent halogen bonding interactions involved in the binding of this synthetic compound to the 80S ribosome.

A single H/ACA small nucleolar RNA mediates tumor suppression downstream of oncogenic RAS

“…The idealization model for tRNA selection traces included four separate FRET values accounting for unbound, initial binding, GTPase activation, and accommodated states during tRNA selection (Ferguson et al, 2015; Geggier et al, 2010) with FRET values of 0.0 ± 0.05, 0.2 ± 0.075, 0.46 ± 0.075 and 0.72 ± 0.075. The idealization model for the pre-translocation state included three FRET states with FRET values of 0.22 ± 0.075, 0.42 ± 0.075 and 0.72 ± 0.075 (0.61 ± 0.05 in case of LD650 labeled tRNA Lys containing ribosomes) accounting for the hybrid 1, hybrid two and classical tRNA binding states (Pellegrino et al, 2019). To generate cumulative distributions for estimation of apparent reaction rates during tRNA selection the number of traces that had achieved the 0.72 FRET state prior to each movie frame were summed.…”

Inhibition of the Eukaryotic 80S Ribosome as a Potential Anticancer Therapy: A Structural Perspective

“…Accessing a wide range of congeners of lissoclimides is key to further investigating the potential of the halogen-π interaction, with the aim to generate more potent and specific drugs. Based on in vitro translational inhibition and IC 50 data [52], the second most potent lissoclimide tested was C45, which contains an additional halogen group in its decalin ring [51]. The crystal structure of the C45/80S complex was of paramount importance to validate previous ab initio docking studies, which suggested a change in conformation of the compound to accommodate into the E-site pocket (Figure 5f).…”

“…It was known that the PET is targeted by a large family of compounds in bacteria, the macrolides; however, recent studies have demonstrated that this region can also be targeted in eukaryotes (Figure 4) [16,20,50]. Finally, the achievement of high-resolution structures of the yeast and human ribosomes allowed unveiling the binding modes of inhibitors targeting the E-site of the LSU, an exclusive eukaryotic-specific feature (Figure 5) [34,51,52].…”

“…Such conformational rearrangements in the PTC would also strongly affect the release of the nascent chain, thereby preventing translation termination to occur. It is very interesting to note the peculiar halogen-π stacking interaction that the chlorine atom of PF846 establishes with the universally conserved A1588 (A752) of 28S rRNA, a rather unusual chemical bonding that recently emerged as a promising method of drug functionalisation [48,51,52,57] (Figure 4b). The aromatic ring where the halogen atom sits additionally forms π-π stacking with the universally conserved U4525 (U2609) (Figure 4b).…”

“…In the same work, the authors compared binding affinities for members of the lissoclimide families to yeast and human translating complexes, with the aim to further characterise the impact of this unusual RNA-ligand interaction. They concluded that the presence of an additional halogen-π interaction may compensate the loss of other types of bondings and, therefore, represent a valuable alternative to provide further stabilisation of the drug within its pocket [51]. Other strategies, together with exploiting the neighbouring eL42 to establish new contacts, would include maintaining CL or CHX scaffolds to design new functional groups that would mimic the A76 of the CCA-end of the deacylated tRNA, which intercalates between G2793 and G2794 during actual translation [51,52,62].…”