S16 throws a conformational switch during assembly of 30S 5′ domain

Rapid and accurate assembly of new ribosomal subunits is essential for cell growth. Here, we show that the ribosomal proteins make assembly more cooperative by discriminating against non-native conformations of the E. coli 16S rRNA. We used hydroxyl radical footprinting to measure how much the proteins stabilize individual rRNA tertiary interactions, revealing the free energy landscape for assembly of the 16S 5′ domain. When ribosomal proteins S4, S17, and S20 bind the 5′ domain RNA, a native and a non-native assembly intermediate are equally populated. The secondary assembly protein S16 suppresses the non-native intermediate, smoothing the path to the native complex. In the final step of 5′ domain assembly, S16 drives a conformational switch at helix 3 that stabilizes pseudoknots in the 30S decoding center. Long-range communication between the S16 binding site and the decoding center helps explain the critical role of S16 in 30S assembly.