Structures of a deAMPylation complex rationalise the switch between antagonistic catalytic activities of FICD

Luke A Perera,Nathan R Zaccai,Steffen Preissler,Michael Haertlein,Sylvain Prévost,David Ron,Juliette M Devos

doi:10.1038/s41467-021-25076-7

Luke A Perera, Nathan R Zaccai + Show 5 more

Open Access

https://doi.org/10.1038/s41467-021-25076-7

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Abstract

The endoplasmic reticulum (ER) Hsp70 chaperone BiP is regulated by AMPylation, a reversible inactivating post-translational modification. Both BiP AMPylation and deAMPylation are catalysed by a single ER-localised enzyme, FICD. Here we present crystallographic and solution structures of a deAMPylation Michaelis complex formed between mammalian AMPylated BiP and FICD. The latter, via its tetratricopeptide repeat domain, binds a surface that is specific to ATP-state Hsp70 chaperones, explaining the exquisite selectivity of FICD for BiP’s ATP-bound conformation both when AMPylating and deAMPylating Thr518. The eukaryotic deAMPylation mechanism thus revealed, rationalises the role of the conserved Fic domain Glu234 as a gatekeeper residue that both inhibits AMPylation and facilitates hydrolytic deAMPylation catalysed by dimeric FICD. These findings point to a monomerisation-induced increase in Glu234 flexibility as the basis of an oligomeric state-dependent switch between FICD’s antagonistic activities, despite a similar mode of engagement of its two substrates — unmodified and AMPylated BiP.

Highlights

The endoplasmic reticulum (ER) Hsp[70] chaperone BiP is regulated by AMPylation, a reversible inactivating post-translational modification
BiP AMPylation inversely correlates with the ER protein folding load, increasing upon the inhibition of protein synthesis[6] and with a resolution of ER stress[3]
The recent discovery that the Enterococcus faecalis Fic protein (EfFic) possesses deAMPylation activity which is dependent on a glutamate homologous to FICD’s Glu23412, suggests conservation of the catalytic mechanism amongst Fic enzymes

Summary

Introduction

The endoplasmic reticulum (ER) Hsp[70] chaperone BiP is regulated by AMPylation, a reversible inactivating post-translational modification. Both BiP AMPylation and deAMPylation are catalysed by a single ER-localised enzyme, FICD. The eukaryotic deAMPylation mechanism revealed, rationalises the role of the conserved Fic domain Glu[234] as a gatekeeper residue that both inhibits AMPylation and facilitates hydrolytic deAMPylation catalysed by dimeric FICD These findings point to a monomerisation-induced increase in Glu[234] flexibility as the basis of an oligomeric statedependent switch between FICD’s antagonistic activities, despite a similar mode of engagement of its two substrates — unmodified and AMPylated BiP. The role of Glu[234] in the oligomeric state-dependent regulation of FICD’s mutually antagonistic activities remains incompletely understood

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