Emerging Quantitative Biochemical, Structural, and Biophysical Methods for Studying Ribosome and Protein-RNA Complex Assembly

Ribosome assembly is one of the most fundamental processes of gene expression and has served as a playground for investigating the molecular mechanisms of how protein-RNA complexes (RNPs) assemble. A bacterial ribosome is composed of around 50 ribosomal proteins, several of which are co-transcriptionally assembled on a similar to 4500-nucleotide-long pre-rRNA transcript that is further processed and modified during transcription, the entire process taking around 2 min in vivo and being assisted by dozens of assembly factors. How this complex molecular process works so efficiently to produce an active ribosome has been investigated over decades, resulting in the development of a plethora of novel approaches that can also be used to study the assembly of other RNPs in prokaryotes and eukaryotes. Here, we review biochemical, structural, and biophysical methods that have been developed and integrated to provide a detailed and quantitative understanding of the complex and intricate molecular process of bacterial ribosome assembly. We also discuss emerging, cutting-edge approaches that could be used in the future to study how transcription, rRNA processing, cellular factors, and the native cellular environment shape ribosome assembly and RNP assembly at large.

Automated summary

Ribosome assembly is a crucial process in gene expression, and studying its molecular mechanisms has led to the development of novel approaches for investigating the assembly of other RNA-protein complexes. This review discusses biochemical, structural, and biophysical methods that have been used to understand the complex process of bacterial ribosome assembly. It also explores emerging approaches for studying how transcription, RNA processing, cellular factors, and the cellular environment influence ribosome and RNA-protein complex assembly.