FIG. 7.
Effect of C-terminal deletions on the self-processing activity of MBP-2793-3143. (A) Total cell lysates from E. coli TB1 cells transformed with pMal-GAV-2793-3143 (lanes 1 and 2; 2793-3143), pMal-GAV-2793-3143_C2968A (lanes 3 and 4; 2793-3143_C2968A), pMal-GAV-2793-3028 (lanes 5 and 6; 2793-3028), and pMal-GAV-2793-3059 (lanes 7 and 8; 2793-3059) were separated by SDS-PAGE in a 12.5% polyacrylamide gel and stained with Coomassie brilliant blue R-250. The bacteria were mock induced (lanes 1, 3, 5, and 7) or induced with 1 mM IPTG for 3 h (lanes 2, 4, 6, and 8). The positions of the fusion proteins and cleavage products are indicated, and the molecular masses of marker proteins (lane M) are given (in kilodaltons). (B) The cell lysates shown in panel A were separated by SDS-PAGE, transferred to a nitrocellulose membrane, and immunostained with anti-MBP antiserum (New England Biolabs). The positions of the uncleaved fusion proteins and the N-terminal (i.e., MBP-containing) cleavage products are indicated, and the positions of marker proteins are given (with masses in kilodaltons).