The ProQ/FinO family of RNA binding proteins mediate sRNA-directed gene regulation throughout gram-negative bacteria. Here, we investigate the structural basis for RNA recognition by ProQ/FinO proteins, through the crystal structure of the ProQ/FinO domain of the Legionella pneumophila DNA uptake regulator, RocC, bound to the transcriptional terminator of its primary partner, the sRNA RocR. The structure reveals specific recognition of the 3’ nucleotide of the terminator by a conserved pocket involving a β-turn-α-helix motif, while the hairpin portion of the terminator is recognized by a conserved α-helical N-cap motif. Structure-guided mutagenesis reveals key RNA contact residues that are critical for RocC/RocR to repress the uptake of environmental DNA in L. pneumophila. Structural analysis and RNA binding studies reveal that other ProQ/FinO domains also recognize related transcriptional terminators with different specificities for the length of the 3’ ssRNA tail.

In prokaryotic species, gene expression is commonly regulated by small, non-coding RNAs (sRNAs). In the gram-negative bacterium Legionella pneumophila, the regulatory, trans-acting sRNA molecule RocR base pairs with a complementary sequence in the 5’-untranslated region of mRNAs encoding for proteins in the bacterial DNA uptake system, thereby controlling natural competence. Sense-antisense duplexing of RocR with targeted mRNAs is mediated by the recently described RNA chaperone RocC. RocC contains a 12 kDa FinO-domain, which acts as sRNA binding platform, along with an extended C-terminal segment that is predicted to be mostly disordered but appears to be required for repression of bacterial competence. In this work we assigned backbone and side chain 1H, 13C, and 15N chemical shifts of RocC’s FinO-domain by solution NMR spectroscopy. The chemical shift data for this protein indicate a mixed α/β fold that is reminiscent of FinO from Escherichia coli. Our NMR resonance assignments provide the basis for a comprehensive analysis of RocC’s chaperoning mechanism on a structural level.