Burridge Charles, Waudby Christopher A, Włodarski Tomasz, Cassaignau Anaïs M E, Cabrita Lisa D, Christodoulou John
Institute of Structural and Molecular Biology, University College London London WC1E 6BT UK
Chem Sci. 2021 Sep 9;12(39):13120-13126. doi: 10.1039/d1sc04313g. eCollection 2021 Oct 13.
The folding of many proteins can begin during biosynthesis on the ribosome and can be modulated by the ribosome itself. Such perturbations are generally believed to be mediated through interactions between the nascent chain and the ribosome surface, but despite recent progress in characterising interactions of unfolded states with the ribosome, and their impact on the initiation of co-translational folding, a complete quantitative analysis of interactions across both folded and unfolded states of a nascent chain has yet to be realised. Here we apply solution-state NMR spectroscopy to measure transverse proton relaxation rates for methyl groups in folded ribosome-nascent chain complexes of the FLN5 filamin domain. We observe substantial increases in relaxation rates for the nascent chain relative to the isolated domain, which can be related to changes in effective rotational correlation times using measurements of relaxation and cross-correlated relaxation in the isolated domain. Using this approach, we can identify interactions between the nascent chain and the ribosome surface, driven predominantly by electrostatics, and by measuring the change in these interactions as the subsequent FLN6 domain emerges, we may deduce their impact on the free energy landscapes associated with the co-translational folding process.
许多蛋白质的折叠可在核糖体上的生物合成过程中开始，并可由核糖体自身进行调节。一般认为，这种干扰是通过新生肽链与核糖体表面之间的相互作用介导的，但是尽管最近在表征未折叠状态与核糖体的相互作用及其对共翻译折叠起始的影响方面取得了进展，但对新生肽链折叠和未折叠状态下的相互作用进行完整的定量分析仍未实现。在这里，我们应用溶液态核磁共振光谱来测量丝状肌动蛋白FLN5结构域的折叠核糖体 - 新生肽链复合物中甲基的横向质子弛豫率。我们观察到，相对于分离的结构域，新生肽链的弛豫率大幅增加，利用分离结构域中的弛豫和交叉弛豫测量，这可能与有效旋转相关时间的变化有关。使用这种方法，我们可以识别主要由静电驱动的新生肽链与核糖体表面之间的相互作用，并且通过测量随着后续FLN6结构域出现时这些相互作用的变化，我们可以推断它们对与共翻译折叠过程相关的自由能景观的影响。
