Fig. 4.

Complementation of pre-AmyE processing in ΔrasP mutant cells. Processing of pre-AmyE (p) to mature AmyE (m) was analyzed by pulse-chase protein labeling with [³⁵S]-methionine in IPTG-induced ΔrasP mutant or wt cells containing either pHTK315-rasP or the empty vector pHTK315. Pre-AmyE and mature AmyE were immunoprecipitated with specific antibodies, separated by LDS-PAGE, and visualized by a phosporimaging. Data from two independent experiments were analyzed with ImageJ to assess the kinetics of pre-AmyE processing to the mature form, and the results are plotted below the autoradiographs. Specifically, the plot shows the relative amounts (%) of the precursor form of AmyE in ΔrasP mutant (triangle) or wt (square) cells carrying pHTK315-rasP (white symbols) or pHTK315 (black symbols) at different time points after the chase with non-radioactive methionine (t = 0)