TOMATO PEROXIDASE: PURIFICATION VIA HYDROPHOBIC CHROMATOGRAPHY

With a large number of isoenzyme species and substrates, peroxidases have been difficult to purify to homogeneity. By including hydrophobic chromatography in the purification scheme, a homogeneous tomato fruit peroxidase isoenzyme was obtained. The isoenzyme had a clean spectrum in the visible region and a R_z (403 nm/280 nm) value of 2.36. It showed up as a single band in disc gel electrophoresis in basic and acidic buffers, in acetate strip electrophoresis, and in both tube and slab gel SDS electrophoresis. Molecular weight estimation of this tomato peroxidase was 43,000 ± 2,000 daltons. The pH optima were at 5.5 and 7.5 with guaiacol and pyrogallol as substrate, respectively. Kinetic studies showed that pyrogallol and hydrogen peroxide could provide substrate inhibition at 5 mM concentration. Hydrophobic chromatography may be useful for purification of other food enzymes similar to tomato peroxidase.