Cat C-specific fluorogenic assay in rat liver lysates.

Abstract

<p><b>A.</b> Labeling of Cat C with FY01. Rat liver extract extracts were treated with increasing concentrations of FY01 for 1 h and labeled proteins analyzed by SDS-PAGE followed by scanning of the gel using a flatbed laser scanner. The location of labeled Cat C is indicated. <b>B.</b> Inhibition of substrate turnover specifically correlates with Cat C labeling. The cleavage of (Pro-Arg)₂-Rho substrate was measured prior to analysis of FY01 labeling shown in part A. Quantification of the indicated labeled proteins relative to DMSO control is shown. <b>C.</b> Cat C-specific HTS assay in rat liver extracts. Rat liver lysates were treated for 30 min with either DMSO or JCP410 (10 µM) followed by the addition of 10 µM of (Pro-Arg)₂-Rho. The turnover rate was continuously measured for 5 min in a 384-well plate. Z’ factor, S/N, and % CV of the negative control are shown.</p