Laboratory of RNA Biophysics and Cellular Physiology, National Heart, Lung and Blood Institute, Bethesda, Maryland 20892-8012, USA.
RNA. 2012 Apr;18(4):759-70. doi: 10.1261/rna.031518.111. Epub 2012 Feb 21.
The archaeal protein L7Ae and eukaryotic homologs such as L30e and 15.5kD comprise the best characterized family of K-turn-binding proteins. K-turns are an RNA motif comprised of a bulge flanked by canonical and noncanonical helices. They are widespread in cellular RNAs, including bacterial gene-regulatory RNAs such as the c-di-GMP-II, lysine, and SAM-I riboswitches, and the T-box. The existence in bacteria of K-turn-binding proteins of the L7Ae family has not been proven, although two hypothetical proteins, YbxF and YlxQ, have been proposed to be L7Ae homologs based on sequence conservation. Using purified, recombinant proteins, we show that Bacillus subtilis YbxF and YlxQ bind K-turns (K(d) ~270 nM and ~2300 nM, respectively). Crystallographic structure determination demonstrates that both YbxF and YlxQ adopt the same overall fold as L7Ae. Unlike the latter, neither bacterial protein recognizes K-loops, a structural motif that lacks the canonical helix of the K-turn. This property is shared between the bacterial and eukaryal family members. Comparison of our structure of YbxF in complex with the K-turn of the SAM-I riboswitch and previously determined structures of archaeal and eukaryal homologs bound to RNA indicates that L7Ae approaches the K-turn at a unique angle, which results in a considerably larger RNA-protein interface dominated by interactions with the noncanonical helix of the K-turn. Thus, the inability of the bacterial and eukaryal L7Ae homologs to bind K-loops probably results from their reliance on interactions with the canonical helix. The biological functions of YbxF and YlxQ remain to be determined.
古菌蛋白 L7Ae 和真核生物同源物，如 L30e 和 15.5kD，构成了最具特征的 K -turn 结合蛋白家族。K-turn 是一种 RNA 模体，由一个凸起组成，两侧是规范和非规范的螺旋。它们广泛存在于细胞 RNA 中，包括细菌基因调控 RNA，如 c-di-GMP-II、赖氨酸和 SAM-I 核糖开关和 T 盒。虽然已经提出了两个假设蛋白 YbxF 和 YlxQ 基于序列保守性是 L7Ae 同源物，但尚未证明细菌中存在 L7Ae 家族的 K-turn 结合蛋白。使用纯化的重组蛋白，我们表明枯草芽孢杆菌 YbxF 和 YlxQ 结合 K-turn（K(d)270 nM 和2300 nM，分别）。晶体结构测定表明，YbxF 和 YlxQ 都采用与 L7Ae 相同的整体折叠。与后者不同，这两种细菌蛋白都不识别 K 环，这是一种缺少 K-turn 规范螺旋的结构模体。这一特性在细菌和真核家族成员之间共享。比较我们的 YbxF 与 SAM-I 核糖开关的 K-turn 复合物的结构和以前确定的与 RNA 结合的古菌和真核同源物的结构表明，L7Ae 以独特的角度接近 K-turn，这导致 RNA-蛋白质界面大得多，主要由与 K-turn 的非规范螺旋的相互作用控制。因此，细菌和真核 L7Ae 同源物不能结合 K 环可能是由于它们依赖于与规范螺旋的相互作用。YbxF 和 YlxQ 的生物学功能仍有待确定。
