Structural basis for dual DNA and RNA specificity of the G-quadruplex-resolving DEAH-box helicase DHX36

DEAH-box helicases, which share a structurally conserved ATPase core, function in all facets of eukaryotic gene expression. While most helicases are highly specialized for their substrates, DHX36 (DEAH-box helicase 36) resolves both DNA and RNA G-quadruplexes. To elucidate the molecular basis of this versatility, we have determined cryo-electron microscopy structures of bovine DHX36 bound to a three-tier RNA G-quadruplex and a six-tier DNA G-quadruplex at 2.6 and 3.4 Å resolution, respectively. Kinetic and smFRET characterizations of structure-guided mutants indicate a key role for the RecA2 domain of the helicase core in DNA vs. RNA discrimination. Furthermore, our structures show that a sequence-divergent RecA2 domain surface loop synergizes with a DHX36-specific N-terminal extension to orthogonally recognize features that specify G-quadruplexes over other nucleic acid structures. Our analysis suggests that recognizing their folded substrates by DEAH-box helicases may generally involve ornamentations of their structural cores acting synergistically with specialized peripheral elements.