Concurrent nucleation of 16S folding and induced fit in 30S ribosome assembly

Publisher Website
Google Scholar
Abstract
Ribosome assembly has to be fast and efficient, given the demand for protein synthesis in rapidly dividing cells. In this work, Woodson and colleagues use a footprinting method with high time resolution to map how the structure of the small subunit's 16S rRNA changes as proteins assemble on it. While several regions of the rRNA adopt secondary structure rapidly, both the region of the decoding site and interactions between the 5′, central and 3′ regions require much more time to form, suggesting that an induced fit reaction is occurring. Rapidly growing cells produce thousands of new ribosomes each minute, in a tightly regulated process that is essential to cell growth1,2. How the Escherichia coli 16S ribosomal RNA and the 20 proteins that make up the 30S ribosomal subunit can assemble correctly in a few minutes remains a challenging problem, partly because of the lack of real-time data on the earliest stages of assembly. By providing snapshots of individual RNA and protein interactions as they emerge in real time, here we show that 30S assembly nucleates concurrently from different points along the rRNA. Time-resolved hydroxyl radical footprinting3 was used to map changes in the structure of the rRNA within 20 milliseconds after the addition of total 30S proteins. Helical junctions in each domain fold within 100 ms. In contrast, interactions surrounding the decoding site and between the 5′, the central and the 3′ domains require 2–200 seconds to form. Unexpectedly, nucleotides contacted by the same protein are protected at different rates, indicating that initial RNA–protein encounter complexes refold during assembly. Although early steps in assembly are linked to intrinsically stable rRNA structure, later steps correspond to regions of induced fit between the proteins and the rRNA.