DEAD-box helicase intrinsically disordered domains and structural dynamics of HIV-1 RNA are required to reveal DDX3X catalytic efficiency.

DEAD-Box helicases are enzymes that bind and remodel RNA and ribonucleoproteins. They are involved in almost every step of RNA metabolism. DEAD-Box helicases are thus major players of gene expression (dys)-regulation and intracellular parasite invasion such as retroviruses. Among many implications in pathologies, the human DEAD-Box helicase DDX3X is hijacked by HIV-1 at various steps including viral RNA export from the nucleus and translation initiation, but little is known about the way it interacts with the viral RNA as well as the structural consequences of this interaction. Here, we show that DDX3X binds to specific regions of HIV-1 5'UTR and dissociates tightly bound dimers of HIV-1 RNA. Such enzymatic activity resulting in the destabilization of a complex structure in multiple turn-over conditions has never been observed with a DEAD-box helicase. DDX3X-induced dynamics was followed using time-resolved structure probing, while footprinting revealed DDX3X preferential binding sites. By coupling the biochemical analysis of DDX3X enzymatic activity the systematic probing of HIV-1-derived RNAs structure, we challenge both the accepted structural model of HIV-1 genomic RNA dimers as well as the dogma considering DEAD box proteins as inefficient and rather promiscuous towards their RNA substrates. An explicative mechanistic model is proposed.