O-acetyltranferases
View Patent ↗A post-translational modification of a predetermined protein expressed by a cell is detected by specifically detecting an O-acetylation of a serine, threonine or tyrosine residue of the protein as an indication of the post-translational modification. The detection may employ antibodies that specifically bind an O-acetylated protein expressed by the cell, wherein the specific binding is dependent on the presence of an O-acetylated serine, threonine or tyrosine residue in the protein.
1. A method of detecting O-acetylation of a substrate protein of an O-acetyltransferase, comprising the step of: specifically detecting O-acetylation of a serine or threonine residue of the substrate protein.
2. The method of claim 1 wherein the specific detecting step comprises contacting the protein with an antibody that specifically binds O-acetylated serine or threonine, and determining that the antibody specifically binds the residue.
3. The method of claim 1 wherein the specific detecting step comprises by mass spectrometry analysis of the protein.
4. The method of claim 1 , wherein the detected O-acetylation occurs under a first physiological condition, and is compared with a control O-acetylation that occurs under a second physiological condition.
5. The method of claim 1 wherein the O-acetylation competes with a different posttranslational modification at the residue, wherein the posttranslational modification is phosphorylation.
6. The method of claim 1 wherein the protein is selected from a MAPK kinase and IκB kinase β (IKKβ).
7. A method of detecting O-acetylation of a substrate protein of an O-acetyltransferase, comprising the steps of:
incubating a mixture comprising the substrate protein, the O-acetyltransferase, and an acetyl donor under conditions wherein the O-acetyltransferase transfers an acetyl from the acetyl donor to a serine or threonine residue of the substrate protein to form the O-acetylation; and specifically detecting the O-acetylation of the serine or threonine residue on the substrate protein.
8. The method of claim 7 wherein the acetyl is radiolabeled.
9. The method of claim 7 wherein the acetyltransferase comprises a catalytic domain of a Clan CE cysteine protease.
10. The method of claim 7 wherein the acetyltransferase comprises a catalytic domain of a Clan CE cysteine protease from a bacterium of a genus selected from the group consisting of Yersinia, Aeromonas, Salmonella, Vibrio, Bartonella, Ralstonia, Pseudomonas, and Xanthomonas.
11. The method of claim 7 wherein the mixture further comprises a candidate agent, and the specifically detected O-acetylation is reduced relative to a control, identifying the candidate agent as an inhibitor of the O-acetyltransferase.
12. The method of claim 7 wherein the mixture further comprises an O-acetyltransferase inhibitor, and the specifically detected O-acetylation is reduced relative to a control.
13. The method of claim 11 or 12 , wherein detecting step comprises specifically detecting a physiological change predetermined to be correlated with the reduced O-acetylation.