EURICA

Post-translational modifications dramatically expand the chemical repertoire of ribosome-synthesised polypeptides and enable otherwise inaccessible molecular functionalities. Some of them are remarkably conserved. For example, the universal ribosomal protein uS11, which in humans is involved in incurable ribosomopathies, undergoes isoaspartylation (i.e. conversion of an asparagine or an aspartate residue into a ß-connected isoaspartate) in most living organisms. Spontaneous isoaspartylation, which kinks the polypeptide chain, is extremely damaging for most proteins and is associated with severe human diseases. However, in uS11 this modification is functionally essential and strictly required for the assembly of the small ribosomal subunit in nearly all species, from E. coli to humans. We identified two families of ancient, deeply conserved enzymes that are most likely responsible for the installation of this unusual modification in bacterial- and eukaryotic/archaeal-type ribosomes. In this ambitious, interdisciplinary project, bringing together experts in bacterial, mitochondrial, eukaryotic RNA biology and ribosome assembly on the one side and those in peptide chemistry and proteomics of post-translational modifications on the other, we will use a wide palette of chemical, biochemical, mass spectrometry, structural, genetic, and physiological approaches to understand the how and why of this unique modification. More specifically, we will undertake a detailed and comprehensive analysis of uS11 isoaspartylation across the tree of life (including model bacteria, archaea, yeast, human cells, protists, and plants). We will decipher the detailed enzymatic mechanisms and idiosyncrasies of this modification. Finally, we will rationalise the essential role of uS11 isoaspartylation in central biological functions of universal significance, including ribosome assembly, protein synthesis, respiration, growth and proliferation.