Small regulatory RNAs (sRNAs) play an important role for posttranscriptional gene regulation in bacteria. sRNAs recognize their target messenger RNAs (mRNAs) by base pairing, which is often facilitated by interactions with the bacterial RNA-binding proteins Hfq or ProQ. The FinO/ProQ RNA-binding protein domain was first discovered in the bacterial repressor of conjugation, FinO. Since then, the functional role of FinO/ProQ-like proteins in posttranscriptional gene regulation was extensively studied in particular in the enterobacteria E. coli and Salmonella enterica and a wide range of sRNA-targets was identified for these proteins. In addition, enterobacterial ProQ homologs also recognize and protect the 3'-ends of a number of mRNAs from exonucleolytic degradation. However, the RNA-binding properties of FinO/ProQ proteins with regard to the recognition of different RNA targets are not yet fully understood. Here, we present the solution NMR structure of the so far functionally uncharacterized ProQ homolog Lpp1663 from Legionella pneumophila as a newly confirmed member and a minimal model system of the FinO/ProQ protein family. In addition, we characterize the RNA-binding preferences of Lpp1663 with high resolution NMR spectroscopy and isothermal titration calorimetry (ITC). Our results suggest a binding preference for single-stranded uridine-rich RNAs in the vicinity of stable stem-loop structures. According to chemical shift perturbation experiments, the single-stranded U-rich RNAs interact mainly with a conserved RNA-binding surface on the concave site of Lpp1663.

中文翻译：

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嗜肺军团菌 (Lpp1663) 的最小 ProQ 同源物的溶液结构和 RNA 结合

小调控 RNA (sRNA) 在细菌转录后基因调控中发挥重要作用。sRNA 通过碱基配对识别其目标信使 RNA (mRNA)，这通常通过与细菌 RNA 结合蛋白 Hfq 或 ProQ 的相互作用来促进。FinO/ProQ RNA 结合蛋白结构域首先在细菌结合抑制因子 FinO 中发现。从那时起，FinO/ProQ 样蛋白在转录后基因调控中的功能作用得到广泛研究，特别是在肠杆菌大肠杆菌和沙门氏菌中，并为这些蛋白质确定了广泛的 sRNA 靶标。此外，肠杆菌 ProQ 同源物还识别并保护许多 mRNA 的 3' 末端免受外切核酸降解。然而，FinO/ProQ 蛋白在识别不同 RNA 靶标方面的 RNA 结合特性尚未完全了解。在这里，我们提出了迄今为止功能未表征的来自嗜肺军团菌的 ProQ 同源物 Lpp1663 的溶液 NMR 结构，作为新确认的成员和 FinO/ProQ 蛋白家族的最小模型系统。此外，我们用高分辨率核磁共振光谱和等温滴定量热法 (ITC) 表征了 Lpp1663 的 RNA 结合偏好。我们的结果表明在稳定的茎环结构附近对单链富含尿苷的 RNA 具有结合偏好。根据化学位移扰动实验，单链富含 U 的 RNA 主要与 Lpp1663 凹位点上的保守 RNA 结合表面相互作用。