Structural basis for the activation of the DEAD-box RNA helicase DbpA by the nascent ribosome

Significance DEAD-box RNA helicases are essential cellular enzymes that remodel misfolded RNA structures in an adenosine triphosphate (ATP)-dependent process. The DEAD-box helicase DbpA is involved in the complex and highly regulated process of ribosome maturation. To prevent wasteful hydrolysis of ATP by DbpA, the enzyme is only active when bound to maturing ribosomes. Here, we elucidate the structural basis behind this important regulatory mechanism and find that the recruited ribosome substrate is able to stabilize the catalytically important closed state of the helicase. In addition, our data identify the natural site of action for DbpA in the maturing ribosome and provide a molecular explanation for the observed ribosome maturation defects that result from the overexpression of a DbpA mutant form.
The adenosine triphosphate (ATP)-dependent DEAD-box RNA helicase DbpA from Escherichia coli functions in ribosome biogenesis. DbpA is targeted to the nascent 50S subunit by an ancillary, carboxyl-terminal RNA recognition motif (RRM) that specifically binds to hairpin 92 (HP92) of the 23S ribosomal RNA (rRNA). The interaction between HP92 and the RRM is required for the helicase activity of the RecA-like core domains of DbpA. Here, we elucidate the structural basis by which DbpA activity is endorsed when the enzyme interacts with the maturing ribosome. We used nuclear magnetic resonance (NMR) spectroscopy to show that the RRM and the carboxyl-terminal RecA-like domain tightly interact. This orients HP92 such that this RNA hairpin can form electrostatic interactions with a positively charged patch in the N-terminal RecA-like domain. Consequently, the enzyme can stably adopt the catalytically important, closed conformation. The substrate binding mode in this complex reveals that a region 5′ to helix 90 in the maturing ribosome is specifically targeted by DbpA. Finally, our results indicate that the ribosome maturation defects induced by a dominant negative DbpA mutation are caused by a delayed dissociation of DbpA from the nascent ribosome. Taken together, our findings provide unique insights into the important regulatory mechanism that modulates the activity of DbpA.