Alternative assembly pathways of ribosomal 30S subunits in thermophilic bacteria
Nathan Napper and Gloria M. Culver
University of Rochester
The ribosome is a large macromolecular complex composed of two asymmetric subunits, each performing critical roles during the translation process. The form and function of this essential cellular machinery has been actively studied, both in vivo and in vitro for several decades. Studies of bacterial 30S subunit assembly have predominantly used E. coli as the model organism. This work has contributed greatly to the knowledge of how 16S rRNA and 20 r- proteins combine to form a functional 30S subunit. It remains unclear, however, if all bacteria use an assembly pathway similar to that of E. coli, or if alternative pathways are used. Bacteria are capable of surviving in diverse and extreme ecological environments, such as extreme temperatures. Due to the temperature dependent nature of 30S subunit assembly in vitro, the way in which thermophilic bacteria assemble ribosomal 30S subunits is of particular interest. To this end we have studied ribosomal 30S subunit assembly in two bacterial species with an optimum growth temperature of 55, Geobacillus stearothermophilus and Geobacillus kaustophilus. Using sucrose gradient sedimentation, tRNA binding, mass spectrometry and 2D gel electrophoresis we have identified and characterized two distinct assembly intermediate populations present in the ribosomal 30S subunit assembly pathway of these bacterial species. These intermediate populations differ in sedimentation properties, function and r-protein composition. This alternative pathway may shed light, both onto the flexibility of assembly of the small ribosomal subunit as well as present novel targets for innovative antibacterial compounds.