In ribosomal RNA precursors the spacer sequences bracketing mature 16 S and 23 S rRNA are base-paired to form long helices (processing stems). In pre-23 S rRNA, the processing stem is continued by eight base-pairs of mature 23 S rRNA known as helix 1. Recently, we have found that any part of 23 S rRNA between positions 40 and 2773 could be deleted without the loss of ribosome-like particle formation, while both end regions were indispensable. In this paper we have analyzed the role of the 5' and 3' end regions of 23 S rRNA during ribosomal 50 S assembly in vivo by using mutants of the 23 S rRNA gene. Deletions and substitutions in both strands of the helix 1 lead to the loss of plasmid derived 50 S formation. Compensatory mutations restoring helix 1 were assembled into functional 50 S subunits. We conclude that the helix 1 of 23 S rRNA is the main RNA determinant for ribosomal large-subunit assembly. Deletions in both the 5' and 3' strand of the processing stem reduced the ability of the 23 S rRNA to form ribosomal 50 S subunits. However, even the complete removal of either the 5' or the 3' strand of the processing stem did not abolish the 50 S assembly completely. Thus, processing stem facilitates, but is not essential for assembly.