Phosphorylation and Methylation of Proteasomal Proteins of the Haloarcheon Haloferax volcanii
Figure 4
Rio1p phosphorylates 1 at Ser and Thr residues. (a) Phosphotransfer of Rio1p to 1 is catalyzed using 1-His6 from recombinant E. coli as a substrate. Phosphotransfer was assayed using the protein substrates indicated as follows: 1-His6 (boiled and unboiled) from recombinant E. coli, 20S CPs of 1-His6 and -StrepII subunits (boiled and unboiled) from H. volcanii and -casein from bovine milk (boiled), as indicated. (b) Phosphotransfer of Rio1p to 1 is reduced by site-directed 1 protein variants Ser58Ala, Thr147Ala and Thr158Ala. The 1 wild type and protein variants were purified from recombinant E. coli and were not boiled prior to assay. Phosphotransfer was detected by autoradiography using Rio1p-StrepII purified by StrepTactin chromatography and P-ATP as a substrate.
