Fig. S8.

Confirmation of TtuB-COOH and TtuB-COSH prepared by the intein method. (A) LC/MS analysis. The samples were reacted with I-AEDANS to stabilize the thiocarboxylate and then analyzed by LC/MS. (Left) Data for the DTT cleavage samples. (Right) Data for the (NH₄)₂S cleavage sample. The extracted ion chromatograms for TtuB-COOH (m/z = 1,045.3, a charge state of 7) and TtuB-COS-AEDANS (m/z = 1,091.4, a charge state of 7) are shown. TtuB-COS-AEDANS was detected only in the (NH₄)₂S cleavage sample. The scanning spectra of TtuB-COOH and TtuB-COS-AEDANS are shown below the chromatograms. The monoisotopic m/z values are closely matched with the theoretical values. (B) Analysis by SDS/PAGE with gels containing APM. TtuB-COOH (lanes 1 and 3) and TtuB-COSH (lanes 2 and 4) prepared by the intein method were analyzed by SDS/PAGE (Left) and SDS/PAGE containing 25 µM APM (Right).