Figure 4.

Pin1 is dose dependently modified by aHNE in MDA-MB-231 cells. (a) Schematic of detection of HNE-adducted proteins in vitro, whereby targets of aHNE are separated from nontargets via click chemistry, followed by streptavidin purification, UV light-induced elution, subject to Western blot, and probed with anti-Pin1 antibody. (b) Western blot of adducted Pin1. Treatment of MDA-MB-231 cells with aHNE and subjection to click-mediated separation of electrophile-modified proteins followed by Western blot with anti-Pin1 antibody reveal a concentration-dependent adduction of Pin1 in this cell line (top blot); the lower blot is shown to demonstrate equal protein loading onto Streptavidin beads.