The Escherichia coli DEAD protein DbpA recognizes a small RNA hairpin in 23S rRNA

The Escherichia coli DEAD protein DbpA is an RNA-specific ATPase that is ac tivated by a 153-nt fragment within domain V of 23S rRNA, A series of RNA s ubfragments and sequence changes were used to identify the recognition elem ents of this RNA-protein interaction. Reducing the size of the fully active 153-nt RNA yields compromised substrates in which both RNA and ATP binding are weakened considerably without affecting the maximal rate of ATP hydrol ysis. All RNAs that stimulate ATPase activity contain hairpin 92 of 23S rRN A, which is known to interact with the 3' end of tRNAs in the ribosomal A-s ite. RNAs with base mutations within this hairpin fail to activate ATP hydr olysis, suggesting that it is a critical recognition element for DbpA. Alth ough the isolated hairpin fails to activate DbpA, RNAs with an extension of approximately 15 nt on either the 5' or 3' side of hairpin 92 elicit full ATPase activity. These results suggest that the binding of DbpA to RNA requ ires sequence-specific interactions with hairpin 92 as well as nonspecific interactions with the RNA extension. A model relating the RNA binding and A TPase activities of DbpA is presented.