New candidate proteins of human liver peroxisomes studied by confocal microscopy.

Human liver peroxisomes were enriched by differential and Nycodenz gradient centrifugation (see Fig. S1). Ten consecutive gradient fractions were quantitatively analyzed by high resolution MS to establish abundance profiles of 314 proteins (Fig. 2, Table S4). Using a supervised statistical approach, the χ2-method, new candidates of human liver peroxisomes were identified with an Euclidian distance (Ρ-value) of ≤1.02. All candidate proteins were shown to associate with human peroxisomes by colocalization studies using confocal microscopy in Huh7 (Fig. 3, Fig. S2 and S6). Sequences of candidate proteins were analyzed for potential peroxisomal targeting signals (PTS) 1 and 2 using both PSORT [31] and the PTS1 predictor [35].