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Magnesium ions are abundant and play indispensable functions in the ribosome. A decrease in Mg2+ concentration causes 70S ribosome dissociation and subsequent unfolding. Structural distortion at low Mg2+ concentrations has been observed in an immature pre50S, while the structural changes in mature subunits have not yet been studied. Here, we purified the 30S subunits of E. coli cells under various Mg2+ concentrations and analyzed their structural distortion by cryo-electron microscopy. Upon systematically interrogating the structural heterogeneity within the 1 mM Mg2+ dataset, we observed 30S particles with different levels of structural distortion in the decoding center, h17, and the 30S head. Our model showed that, when the Mg2+ concentration decreases, the decoding center distorts, starting from h44 and followed by the shifting of h18 and h27, as well as the dissociation of ribosomal protein S12. Mg2+ deficiency also eliminates the interactions between h17, h10, h15, and S16, resulting in the movement of h17 towards the tip of h6. More flexible structures were observed in the 30S head and platform, showing high variability in these regions. In summary, the structures resolved here showed several prominent distortion events in the decoding center and h17. The requirement for Mg2+ in ribosomes suggests that the conformational changes reported here are likely shared due to a lack of cellular Mg2+ in all domains of life.

Myo2, a yeast class V myosin, transports a broad range of organelles and plays important roles in various cellular processes, including cell division in budding yeast. Despite the fact that several structures of Myo2/cargo adaptor complexes have been determined, the understanding of the versatile cargo-binding modes of Myo2 is still very limited, given the large number of cargo adaptors identified for Myo2. Here, we used ColabFold, an AlphaFold2-powered and easy-to-use tool, to predict the complex structures of Myo2-GTD and its several cargo adaptors. After benchmarking the prediction strategy with three Myo2/cargo adaptor complexes that have been determined previously, we successfully predicted the atomic structures of Myo2-GTD in complex with another three cargo adaptors, Vac17, Kar9 and Pea2, which were confirmed by our biochemical characterizations. By systematically comparing the interaction details of the six complexes of Myo2 and its cargo adaptors, we summarized the cargo-binding modes on the three conserved sites of Myo2-GTD, providing an overall picture of the versatile cargo-recognition mechanisms of Myo2. In addition, our study demonstrates an efficient and effective solution to study protein–protein interactions in the future via the AlphaFold2-powered prediction.