Formation of 100S ribosome dimer is generally associated with translation suppression in bacteria. Trans-acting factors RMF and HPF were shown to directly mediate this process in E. coli. Gram-positive S. aureus lacks an RMF homologue, and the structural basis for its 100S formation was not known. Here we report the cryo-EM structure of the native 100S ribosome from S. aureus, revealing the molecular mechanism of its formation. The structure is distinct from previously reported analogues and relies on the HPF C-terminal extension forming the binding platform for the interactions between both of the small ribosomal subunits. The 100S dimer is formed through interactions between rRNA h26, h40 and protein uS2, involving conformational changes of the head as well as surface regions that could potentially prevent RNA polymerase from docking to the ribosome.
