Insights into G-quadruplex specific recognition by the DEAH-box helicase RHAU: Solution structure of a peptide–quadruplex complex

Abstract

Four-stranded nucleic acid structures called G-quadruplexes have been associated with important cellular processes, which should require G-quadruplex-protein interaction. However, the structural basis for specific G-quadruplex recognition by proteins has not been understood. The DEAH (Asp-Glu-Ala-His) box RNA helicase associated with AU-rich element (RHAU) (also named DHX36 or G4R1) specifically binds to and resolves parallel-stranded G-quadruplexes. Here we identified an 18-amino acid G-quadruplex-binding domain of RHAU and determined the structure of this peptide bound to a parallel DNA G-quadruplex. Our structure explains how RHAU specifically recognizes parallel G-quadruplexes. The peptide covers a terminal guanine base tetrad (G-tetrad), and clamps the G-quadruplex using three-anchor-point electrostatic interactions between three positively charged amino acids and negatively charged phosphate groups. This binding mode is strikingly similar to that of most ligands selected for specific G-quadruplex targeting. Binding to an exposed G-tetrad represents a simple and efficient way to specifically target G-quadruplex structures.G-quadruplex | RHAU helicase | DHX36 | DEAH-box family | NMR

Oligomer formation and G-quadruplex binding by purified murine Rif1 protein, a key organizer of higher-order chromatin architecture

“…Thus, muRif1 shows some preference for G4 structures with particular topology (parallel, hybrid) over other topology (anti-parallel, telomere-derived). Then, we focused on parallel-type G4 oligonucleotides consisting of different nucleotide sequences (47), and evaluated their binding to the full-length muRif1 in EMSA (Fig. 7A).…”

The G-quadruplex (G4) resolvase DHX36 efficiently and specifically disrupts DNA G4s via a translocation-based helicase mechanism

“…The disruption rate was shown to be greater for a G4 with fewer G-quartets, suggesting that less stable G4s are disrupted more efficiently by DHX36 (26). A small Nterminal domain directs DHX36 to G4s by binding specifically to these structures (28)(29)(30).…”

“…Tetramolecular G4s are attractive models for monitoring DHX36 activity because they predominantly adopt the parallel conformation (26,35,36), which is preferentially recognized by the Nterminal domain of DHX36 (30). In addition, tetramolecular G4s form slowly (37), allowing their disruption to be monitored by an electrophoretic mobility shift assay (EMSA) without the need for a 'chase' oligonucleotide to prevent G4 reformation (14,26,38) (Fig.…”

“…This preference is presumably due at least in part to the N-terminal domain of DHX36, which binds directly to G4s and is thought to direct DHX36 to these structures (29,30).…”

“…The disruption rate was shown to be greater for a G4 with fewer G-quartets, suggesting that less stable G4s are disrupted more efficiently by DHX36 (26). A small Nterminal domain directs DHX36 to G4s by binding specifically to these structures (28)(29)(30).Nevertheless, important aspects of the mechanism of G4 disruption by DHX36 remain unknown. DEAH/RHA family helicases (abbreviated herein as DEAH family) typically use a translocation-based mechanism for unwinding DNA or RNA helices, a mechanism in which ATP drives directional motion along a sequestered single strand, displacing the complementary strand (31).…”