AMPylation: Something Old is New Again

Abstract

The post-translational modification AMPylation is emerging as a significant regulatory mechanism in both prokaryotic and eukaryotic biology. This process involves the covalent addition of an adenosine monophosphate to a protein resulting in a modified protein with altered activity. Proteins capable of catalyzing AMPylation, termed AMPylators, are comparable to kinases in that they both hydrolyze ATP and reversibly transfer a part of this primary metabolite to a hydroxyl side chain of the protein substrate. To date, only four AMPylators have been characterized, though many more potential candidates have been identified through amino acid sequence analysis and preliminary in vitro studies. This modification was first discovered over 40 years ago by Earl Stadtman and colleagues through the modification of glutamine synthetase by adenylyl transferase; however research into this mechanism has only just been reenergized by the studies on bacterial effectors. New AMPylators were revealed due to the discovery that a bacterial effector having a conserved Fic domain transfers an AMP group to protein substrates. Current research focuses on identifying and characterizing various types of AMPylators homologous to Fic domains and adenylyl transferase domains and their respective substrates. While all AMPylators characterized thus far are bacterial proteins, the conservation of the Fic domain in eukaryotic organisms suggests that AMPylation is omnipresent in various forms of life and has significant impact on a wide range of regulatory processes.

Keywords: AMPylation; Fic; GTPase; Type III secretion system; adenylyl transferase; adenylylation; effector.

Figure 1

AMPylation in pathogenicity. (A) The Fic effector AMPylators VopS (shown) and IbpA are secreted into eukaryotic cells and modify a threonine or tyrosine residue on the Switch 1 loop of Rho family GTPases, sterically blocking their association with downstream substrates like PAK and rhotekin. (B) The adenylyl transferase effector AMPylator DrrA catalyzes the exchange of GDP for GTP in the GTPase Rab1b through its GEF domain (not shown) and AMPylates a tyrosine in the Switch 2 loop, preventing its association with GAPs. (C, D) HeLa cells transfected with mock vector (C) or with VopS (D), visualized with confocal microscopy for rhodamine phalloidin stain (actin, red) and Hoechst stain (DNA, blue).

Figure 2

Domain architecture of the Fic-containing protein VopS and the adenylyl transferase region of DrrA. The Fic domain is highlighted in red with the non-conserved subdomain in white. Black spheres in VopS display the conserved catalytic histidine of the Fic motif. The DrrA adenylyl transferase region is colored green and black spheres correspond to the two conserved aspartates in adenylyl transferases.