The Native 3D Organization of Bacterial Polysomes

Recent advances have led to insights into the struc-ture of the bacterial ribosome, but little is knownabout the 3D organization of ribosomes in the context of translating polysomes. We employed cryoelectron tomography and a template-matching approach to map 70S ribosomes in vitrified bacterial translation extracts and in lysates of active E. coli spheroplasts. In these preparations, polysomal arrangements were observed inwhich neighboring ribosomesare densely packed and exhibit preferred orientations. Analysis of characteristic examples of polysomes reveals a stag-gered or pseudohelical organization of ribosomes along the mRNA trace, with the transcript being sequestered on the inside, the tRNA entrance sites being accessible, and thepolypeptide exit sites facing the cytosol. Modeling of elongating nascent polypep-tide chains suggests that this arrangementmaximizes the distance between nascent chains on adjacent ribosomes, thereby reducing the probability of inter-molecular interactions thatwouldgive rise toaggrega-tion and limit productive folding