Elongation factor G (EF-G) catalyzes ribosome’s fast and accurate translocation on the mRNA. Although the high-resolution structures of EF-G bound to the ribosome have been determined by cryo-EM and X-ray studies, the correlation between the structure and the EF-G generated power stroke force remains elusive. It is also highly debated regarding the amplitude of the EF-G power stroke. In the present work, we report the synthesis of a series of crosslinked EF-G’s and the precise measurements of their power stroke. The crosslinked EF-G’s contained different lengths of linker molecules, which restricted the conformational changes to different extents. The power strokes were determined by an atomic magnetometer or a microscope. Both methods yielded consistent results. The results showed that crosslinked EF-G with a short linker generated much smaller power stroke compared to the 89 pN force that was generated by the wild type EF-G, whereas a longer link did not significantly affect the power stroke amplitude. We also investigated the correlation between power stroke amplitude and translocation efficiency. It showed that the smaller power stroke reduced the translocation rate but not the step size. Furthermore, our research provides a widely adoptable technique for measuring protein power stroke.

Visualization and modulation of EF-G power stroke in ribosomal translocation

2019 Spring National Meeting

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