Generation of Mature Nα-Terminal Acetylated Thymosin α1 by Cleavage of Recombinant Prothymosin α
Figure 2
(a) Expression and purification of legumain in P. pastoris and its autoactivation. SDS-PAGE of autocatalytic legumain after the prolegumain was incubated in buffer as a function of time (0–4 h) at 37°C. Aliquots of each reaction were analyzed by SDS-PAGE (12%) and stained with Coomassie blue. Each time point is marked on the bottom of its corresponding lane. (b) SDS-PAGE analysis of Prothymosin α proteolysis by recombinant legumain in vitro. Aliquots of ProTα were incubated with recombinant legumain at 37°C in buffer containing 1 mM DTT, 1 mM EDTA, and 0.1 M sodium citrate (pH 4.0). Each reaction mixture was analyzed by SDS-PAGE under the conditions indicated under “Experimental Section.” Each time point is marked on the bottom of its corresponding lane 1–6. Lane 7 is the chemically synthesized Nα-acetylated Tα1.
