Catalytic characterization of heterodimeric linoleate 13S-lipoxygenase from black soybean (Glycine max (L.) Merr.).

A novel lipoxygenase (BLOX) was purified from black soybean (Glycine max (L.) Merr.), and its catalytic properties were characterized. The molecular weight of BLOX was 101 kDa and its unique heterodimeric structure with two different subunits of molecular weight 46 kDa and 55 kDa was elucidated. The optimum pH and temperature of BLOX were pH 9.5 and 40 °C, respectively. BLOX was highly stable at the pH range of 6.0-10.0 and below 40 °C, and was stimulated by adding ferrous ion (Fe2+). In terms of substrate specificity, BLOX showed a substrate preference to linoleic acid that is the main substance to produce hydroperoxides in soybean. When it reacted with linoleic acid, the major product was 13(S)-hydroperoxy-9,11-octadecadienoic acid; therefore, it could be classified into the linoleate 13S-LOX family (EC 1.13.11.12). Finally, the kinetic parameters (Vmax, Km, and kcat) of BLOX were 0.124 mM min-1, 0.636 mM, and 12.28 s-1, respectively, and consequently, the catalytic efficiency (kcat/Km) was calculated as 1.93 × 104 M-1·s-1. These catalytic characteristics of BLOX could contribute to understanding the enzymatic rancidification of black soybean, and to further biotechnical approaches to control and mitigate the deterioration.