Analysis of DegU protein variants produced
(A) Western blot analysis of proteins extracted from 3610 (wild-type strain), degU (NRS1314), degU + P
-degU (H12L)-lacI (NRS1326), degU + P
-degU (D56N)-lacI (NRS1327), degU + Phy-spank-degU
(H12L)-lacI (NRS1325) and degU + P
-degU (H12L, D56N)-lacI (NRS4762), grown in the absence (−) or presence (+) of 50 μM IPTG. The band corresponding to DegU is indicated by an asterisk. The blot was probed with a DegU-specific polyclonal antibody. (B) Coomassie stained SDS-PAGE analysis of purified recombinant proteins. Shown are: DegU WT (pNW43), DegU D56N (pNW52), DegU H12L (pNW54) and DegU H12L D56N (pNW1072). (C) Circular dichroism analysis of the proteins detailed in (B). The plotted spectra are normalised to each other to account for concentration and zeroed at 260 nm after the background spectrum was subtracted.