Buskiewicz Iwona A, Jöckel Johannes, Rodnina Marina V, Wintermeyer Wolfgang
Institute of Molecular Biology, University of Witten/Herdecke, 58448 Witten, Germany.
RNA. 2009 Jan;15(1):44-54. doi: 10.1261/rna.1285609. Epub 2008 Nov 24.
The bacterial signal recognition particle (SRP) binds to ribosomes synthesizing inner membrane proteins and, by interaction with the SRP receptor, FtsY, targets them to the translocon at the membrane. Here we probe the conformation of SRP and SRP protein, Ffh, at different stages of targeting by measuring fluorescence resonance energy transfer (FRET) between fluorophores placed at various positions within SRP. Distances derived from FRET indicate that SRP binding to nontranslating ribosomes triggers a global conformational change of SRP that facilitates binding of the SRP receptor, FtsY. Binding of SRP to a signal-anchor sequence exposed on a ribosome-nascent chain complex (RNC) causes a further change of the SRP conformation, involving the flexible part of the Ffh(M) domain, which increases the affinity for FtsY of ribosome-bound SRP up to the affinity exhibited by the isolated NG domain of Ffh. This indicates that in the RNC-SRP complex the Ffh(NG) domain is fully exposed for binding FtsY to form the targeting complex. Binding of FtsY to the RNC-SRP complex results in a limited conformational change of SRP, which may initiate subsequent targeting steps.
细菌信号识别颗粒（SRP）与正在合成内膜蛋白的核糖体结合，并通过与SRP受体FtsY相互作用，将它们靶向到膜上的转运体。在这里，我们通过测量位于SRP内不同位置的荧光团之间的荧光共振能量转移（FRET），来探究SRP和SRP蛋白Ffh在靶向不同阶段的构象。从FRET得出的距离表明，SRP与非翻译核糖体的结合触发了SRP的整体构象变化，这有利于SRP受体FtsY的结合。SRP与核糖体-新生链复合物（RNC）上暴露的信号锚定序列的结合会导致SRP构象的进一步变化，涉及Ffh（M）结构域的柔性部分，这将核糖体结合的SRP对FtsY的亲和力提高到Ffh分离的NG结构域所表现出的亲和力。这表明在RNC-SRP复合物中，Ffh（NG）结构域完全暴露以结合FtsY形成靶向复合物。FtsY与RNC-SRP复合物的结合导致SRP的构象变化有限，这可能启动后续的靶向步骤。
