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. Author manuscript; available in PMC: 2012 Nov 2.
Published in final edited form as: J Am Chem Soc. 2011 Oct 7;133(43):17103–17105. doi: 10.1021/ja205137d

Figure 2.

Figure 2

Detection of endogenous AMPylation substrates in cell lysates and proteomic substrate identification. AMPylation reactions (A–C) were carried out in 15 μl for 1 h at 30°C A) VopS (100 ng) was added to HeLa cell lysate (10 mg) in the presence of N6pATP (100 μM). B) Fic2 (300 ng) was added to HeLa cell lysate (10 mg) in the presence of N6pATP (100 μM). C) DrrA (4 μg) was added to HeLa cell lysate (10 mg) in the presence of N6pATP (100 μM). D) Proteomic analysis of VopS substrates in HeLa cell lysate revealed two protein IDs above background, VopS and Cdc42. E) Identified Cdc42 peptides. # TS – number of total spectra, # UP – number of unique peptides, XCorr – SEQUEST Xcorr score, DCn – SEQUEST ΔCn score, az-rho - azido-rhodamine fluorescence; CB, coomassie blue; asterisk indicates auto-AMPylation and recombinant AMPylator.