F-actin sequesters elongation factor 1α from interaction with aminoacyl-tRNA in a pH-dependent reaction

Abstract

The machinery of eukaryotic protein synthesis is found in association with the actin cytoskeleton. A major component of this translational apparatus, which is involved in the shuttling of aa-tRNA, is the actin- binding protein elongation factor 1α (EF-1α). To investigate the consequences for translation of the interaction of EF-1α with F-actin, we have studied the effect of F-actin on the ability of EF-1α to bind to aa- tRNA. We demonstrate that binding of EF-1α:GTP to aa-tRNA is not pH sensitive with a constant binding affinity of ~0.2 μM over the physiological range of pH. However, the sharp pH dependence of binding of EF- 1α to F-actin is sufficient to shift the binding of EF-1α from F-actin to aa-tRNA as pH increases. The ability of EF-1α to bind either F-actin or aa- tRNA in competition binding experiments is also consistent with the observation that EF-1α's binding to F-actin and aa-tRNA is mutually exclusive. Two pH-sensitive actin-binding sequences in EF-1α are identified and are predicted to overlap with the aa-tRNA-binding sites. Our results suggest that pH-regulated recruitment and release of EF-1α from actin filaments in vivo will supply a high local concentration of EF-1α to facilitate polypeptide elongation by the F-actin-associated translational apparatus.