Kinetics of soybean lipoxygenases are related to pH, substrate availability and extraction procedures

The kinetic patterns of pure soy lipoxygenase LOX-1 and crude or defatted soybean extracts containing LOX isoenzymes (LOX-1, LOX-2 and LOX-3) were studied by UV spectrometry at 234 and 280 nm, depending on their extraction and measurement conditions. Different pHs (from 6.0 to 9.0), corresponding to specific activation of LOX isoenzymes and the ratios of enzyme protein per substrate were used in order to evaluate the enzyme rates, as indicators of its affinity for substrate in different environments. The crude soy extract contained mainly LOX-1 activity (measured at 234 nm, at pH 9.0) and LOX-3, in an approximate ratio of 3:1. The LOX-2 activity was very low. The defatted extracts buffered at pH 6.8 and 7.1 showed a low LOX-1 and LOX 2 activity, but mostly LOX-3 activity (measured at 280 nm, at pH 7.1), with a mirror-type relation between the enzyme/substrate ratio and their enzymatic specific activity. The results suggest that defatting inhibits specifically the LOX-1 activity and indicate the possibility to modulate LOX activity by modifications of enzyme/substrate ratios and modifications of pH in the enzyme environment.