Figure 3

TPP distribution in a light mitochondrial fraction after Nycodenz gradient centrifugation. The light mitochondrial fractions obtained from rat liver were subjected to Nycodenz gradient centrifugation. Fractions were collected starting from the bottom. In a first experiment (panel A), TPP content (black line) and catalase activity (marker enzyme for the peroxisomal matrix; white bars) were measured. Recoveries for TPP content and catalase activity were 60% and 70%, respectively. Thiamine, which is also present in the L-fraction, was recovered in the top fractions (data not shown). The insert shows the density (g/ml) of the gradient. In a second experiment (panels B-D), HACL1 activity was measured (panel B; white bars; recovery 78%) revealing the highest activity (35 mU/mg protein) in fraction 6, corresponding to the TPP peak (180 pmol/mg protein, black line). In panels C and D, the distribution of LDH (marker for cytosol; recovery 121%) and urate oxidase (marker for peroxisomal core; recovery 66%) is presented (white bars).