The C-terminal domain of yeast Arginyltransferase1 is essential for its catalytic activity.

Arginyltransferase 1 (Ate1), a conserved eukaryotic enzyme, has recently been structurally characterised; identifying a conserved N-terminal domain, a central GNAT fold and a variable C-terminal domain of unknown function. Here, using budding yeast as an experimental model, we established that the C-terminal domain is indispensable for Ate1 stability, its enzymatic activity and Ate1-mediated cell death. Furthermore, we found that the co-expression of separated N-terminal and C-terminal halves of Ate1 did not reconstitute enzyme activity in vivo. Also, we observed that the mutation of conserved residues involved in cofactor binding and active site formation within the N-terminal half of Ate1 abolishes the enzyme function. Moreover, we showed that mouse Ate1, when heterogeneously expressed in yeast, was catalytically active but not lethal. These findings underscore the structural interdependence of Ate1 domains and their integrity in maintaining enzyme catalytic activity and stability, providing deeper insights into the enzyme structure-function relationship.