Multi-protein assemblies orchestrate co-translational enzymatic processing on the human ribosome

Most nascent chains undergo rapid co-translational enzymatic processing as soon as their N-terminus becomes accessible at the ribosomal polypeptide tunnel exit (PTE). In eukaryotes, N-terminal methionine excision (NME) by Methionine Aminopeptidases (MAP1 and MAP2), and N-terminal acetylation (NTA) by N-Acetyl-Transferase A (NatA), is the most common set of subsequent modifications carried out on the ribosome. How these two enzymatic processes are coordinated in quick succession in the context of a rapidly translating ribosome has remained elusive. Here, we report that human NatA occupies a non-intrusive distal binding site on the ribosome which does not interfere with most other ribosome associated factors (RAFs). In this position, NatA can partake in a coordinated and dynamic assembly with MAP1 through a complex scaffolding function of the abundant Nascent Polypeptide Associated Complex (NAC). Alternatively, MAP2 can co-occupy the PTE with NatA in preparation for successive NME and NTA. In contrast to MAP1, MAP2 completely covers the PTE and is thus incompatible with NAC and MAP1 recruitment. Both assemblies can compile on the human ribosome independent of nascent chain substrates. Together, our structures provide the structural framework for the rapid coordinated orchestration of NME and NTA in protein biogenesis.