Structural basis for EarP-mediated arginine glycosylation of translation elongation factor EF-P

Abstract

15Glycosylation is a universal strategy to post-translationally modify proteins. The recently discovered 16 arginine rhamnosylation activates the polyproline specific bacterial translation elongation factor EF-P. 17 EF-P is rhamnosylated on arginine 32 by the glycosyltransferase EarP. However, the enzymatic 18 mechanism remains elusive. In the present study, we solved the crystal structure of EarP from 19Pseudomonas putida. The enzyme is composed of two opposing domains with Rossmann-folds, thus 20 constituting a GT-B glycosyltransferase. While TDP-rhamnose is located within a highly conserved 21 pocket of the C-domain, EarP recognizes EF-P via its KOW-like N-domain. Based on our structural 22 data combined with an in vitro / in vivo enzyme characterization, we propose a mechanism of 23 inverting arginine glycosylation. As EarP is essential for pathogenicity in P. aeruginosa our study 24 provides the basis for targeted inhibitor design. 25