Adenylylation

Adenylylation,[1][2] now known as AMPylation, is a process in which adenosine monophosphate (AMP) molecule is covalently attached to a protein side chain, altering the function of the protein.[3] This covalent addition of AMP to a hydroxyl side chain of the protein is posttranslational modification that is stable and reversible.[4] Adenylylation involves a phosphodiester bond between a hydroxyl group of the molecule undergoing adenylylation and the phosphate group of the adenosine monophosphate nucleotide (i.e. adenylic acid). This process can occur to molecules such as tyrosine residues. Enzymes that are capable of catalyzing this process are called AMPylators.

Function

Similar to serine, threonine or tyrosine phosphorylation, AMPylation regulates the activity of some proteins, such as glutamine synthetase.[5] Additionally, AMPylation aids in allowing thermodynamically unfavorable overall reactions to take place by generating a leaving group in chemical mechanisms that indirectly use energy from ATP hydrolysis.[6] Unstable carboxylate-phosphate mixed anhydrides or of phosphoramidates are generated in this transient adenylyation reaction[3]

The degree of adenylylation depends on the ratio of glutamine to α-ketoglutarate: The higher this ratio the more monomers are adenylylated, thereby producing lower activity of glutamine synthetase; the lower the ratio the less monomers are adenylated and the higher activity of glutamine synthetase. A high ratio is a sign of cellular nitrogen sufficiency, whereas a low ratio is evidence of a limited nitrogen and the need for ammonia fixation by glutamine synthetase.[2]

AMPylators

AMPylators are enzymes that catalyze AMPylation. These enzymes have been shown to be comparable to kinases due to their ATP hydrolysis activity and reversible transfer of the metabolite to a hydroxyl side chain of the protein substrate. To date, the AMPylators that have been identified are bacterial proteins. Two domains, the Fic and adenylyl transferase domains, are the currently known AMPylators that have been shown to be involved in pathogenicity of bacterial species and metabolic regulation. Fic domains are evolutionarily conserved domains in prokaryotes and eukaryotes that belong to the Fido domain superfamily, whereas the adenylyl transferase domains are part of the larger nucleotidyl transferase protein family[4]

GTPases are common targets of AMPylators. Rho, Rab, and Arf GTPase families are involved in actin cytoskeleton dynamics and vesicular trafficking. They also play roles in cellular control mechanisms such as phagocytosis in the host cell, in which the pathogen enhances or prevents its internalization by either inducing or inhibiting host cell phagocytosis[4]

AMPylation and pathogenicity

Bacteria proteins, also known as effectors, have been shown to use AMPylation. Effectors such as VopS, IbpA, and DrrA, have been shown to AMPylate host GTPases and cause actin cytoskeleton changes.

Vibrio parahaemolyticus or VopS is a Gram-negative bacterium that causes food poisoning as a result of raw or undercooked seafood consumption in humans.[7] VopS contains a Fic domain that has a conserved HPFx(D/E)GN(G/K)R motif that contains a histidine residue essential for AMPylation. VopS blocks actin assembly by modifying threonine residue in the switch 1 region of Rho GTPases. The transfer of an AMP moiety using ATP to the threonine residue results in steric hindrance, and thus prevents Rho GTPases from interacting with downstream effectors. As a result, the host cell’s actin cytoskeleton control is disabled, leading to cell rounding.[4][7]

IbpA is secreted into eukaryotic cells from H. somni, a Gram-negative bacterium in cattle that causes respiratory epithelium infection. This effector contains two Fic domains at the C-terminal region. AMPylation of the IbpA Fic domain of Rho family GTPases is responsible for its cytotoxicity. The AMPylation on a tyrosine residue of the switch 1 region blocks the interaction of the GTPases with downstream substrates such as PAK.

De-AMPylation

GS-ATasE (GlnE) is an AMPylator that has been shown to catalyze de-AMPylation of glutamine synthetase by removing the covalent linkage between AMP and a hydroxyl residue of a protein. It contains two adenylyl transferase domains that are involved in the addition and removal of AMP to glutamine synthetase. De-AMPylation occurs at the N-terminus of the domain. Following the removal of AMP from glutamine synthetase, GS-ATase forms ADP and unmodified glutamine synthetase.[4]

References

  1. ↑ Han KK, Martinage A (1992). "Post-translational chemical modification(s) of proteins". Int. J. Biochem. 24 (1): 19–28. PMID 1582530.
  2. 1 2 Garrett, R.H., and C.M. Grisham. Biochemistry. 3rd ed. Belmont, CA: Thomas, 2007. 815-20
  3. 1 2 Itzen, Aymelt, Wulf Blankenfeldt, and Roger S. Goody. "Adenylylation: renaissance of a forgotten post-translational modification." Trends in Biochemical Sciences 36.4 (2011): 221-228. Print.
  4. 1 2 3 4 5 Woolery, Andrew. "AMPylation: something old is new again." Frontiers in Microbiology 1 (2010): 1-18. Print.
  5. ↑ Stadtman, E.R. (2001) The story of glutamine synthetase regulation. J. Biol. Chem. 276, 44357–44364
  6. ↑ Schmelz, S. and Naismith, J.H. (2009) Adenylate-forming enzymes. Curr. Opin. Struct. Biol. 19, 666–671
  7. 1 2 Luong, P., L. N. Kinch, C. A. Brautigam, N. V. Grishin, D. R. Tomchick, and K. Orth. "Kinetic and Structural Insights into the Mechanism of AMPylation by VopS Fic Domain." Journal of Biological Chemistry 285.26 (2010): 20155-20163. Print.


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