AMPylation – a new use for ATP
Post-translational modifications are key steps in the regulation of eukaryotic cell signaling. The modification of a protein can enhance or change the role of the protein. For instance, when a protein is phosphorylated, it might become an active enzyme, be targeted to a subcellular compartment or be efficiently degraded. In addition, the modified protein could act as a scaffold to cluster proteins and form signaling networks.
We recently discovered a bacterial virulence factor from Vibrio parahaemolyticus that mediates the modification of a conserved threonine residue on eukaryotic substrates via a phosphodiester bond with AMP (Yarbrough et al., Science 2009). This modification, called AMPylation, is mediated by a domain called a filamentation induced by cAMP (Fic) domain. We have solved the structure of VopS Fic domain and characterized the kinetics of AMPylation (Luong et al., J Biol Chem 2010).
As Fic domains are evolutionarily conserved in bacteria as well as higher organisms and evidence has been found for AMPylation occurring in lysates from human tissue culture cells, we are currently studying the role of this post-translational modification in eukaryotes. To accomplish this, we are developing tools to detect specific AMPylation, including an antibody against AMPylated threonine (Hao et al., J Biotechnol 2011).
Publications on AMPylation
- Ham H., Woolery A.R., Tracy C., Stenesen D., Krämer H. & Orth, K. Unfolded protein response-regulated dFic reversibly AMPylates BiP during ER homeostasis. (2014) JBC (In Press).
- Yu, X., Woolery, A.R., Luong, P., Hao, Y.H., Grammel, M., Westcott, N., Park, J., Wang, J., Bian, X., Demirkan, G., Hang, H.C., Orth, K. & LaBaer, J. Click chemistry-based detection of global pathogen-host AMPylation on self-assembled human protein microarrays. (2014) Mol & Cell Proteomics (In Press).
- Lewallen,D.M., Sreelatha, A., Dharmarajan, V., Madoux, F., Chase,, F. ,Griffin, P.R., Orth, K., Hoddler, P. & Thompson, P.R. Inhibiting AMPylation: a novel screen to identify the first small molecule inhibitors of protein AMPylation. ACS Chem Biol (2014) 9(2): 433-432 PMC3944102.
- Ham H & Orth, K. De-AMPylation unmasked: modulation of host membrane trafficking. Sci Signal. (2011) Oct 11;4(194).
- Grammel M, Luong P, Orth, K., Hang HC. A chemical reporter for protein AMPylation. J Am Chem Soc. (2011) Nov 2;133(43):17103-5.
- Hao YH, Chuang T, Ball HL, Luong P, Li Y, Flores-Saaib RD, Orth, K. Characterization of a rabbit polyclonal antibody against threonine-AMPylation. (2011) J Biotechnol. 151(3):251-4.
- Li Y, Al-Eryani R, Yerbrough ML, Orth, K., Ball HL. Characterization of AMPylation on threonine, serine, and tyrosine using mass spectrometry. (2011) J Am Soc Mass Spectrom. 22(4):752-61.
- Luong P, Kinch LN, Brautigam CA, Grishin NV, Tomchick DR, Orth, K. Kinetic and structural insights into the mechanism of AMPylation by VopS Fic domain. (2010) J Biol Chem. 285(26):20155-63.
- Woolery AR, Luong P, Broberg CA, Orth, K. AMPylation: Something Old is New Again. (2010) Front Microbiol. 1:113.
- Kinch LN, Yarbrough ML, Orth, K., Grishin NV. Fido, a novel AMPylation domain common to fic, doc, and AvrB. (2009) PLoS One. 4(6):e5818.
- Yarbrough ML, Orth, K. AMPylation is a new post-translational modification. (2009) Nat Chem Biol. 5(6):378-9.
- Yarbrough ML, Li Y, Kinch LN, Grishin NV, Ball HL, Orth, K. AMPylation of Rho GTPases by Vibrio VopS disrupts effector binding and downstream signaling. (2009) Science. 323(5911):269-72.