Insight into naphthoquinone metabolism:: β-glucosidase-catalysed hydrolysis of hydrojuglone β-D-glucopyranoside

In plants, the naphthoquinone juglone is known to be involved in pathogenic defence mechanisms, but it may also take part in plant developmental processes. This naphthoquinone can accumulate in a glycosylated form, namely hydrojuglone beta-D-glucopyranoside. The structural configuration of this compound was shown to be 1,5-dihydroxy-4-naphthalenyl-beta-D-glucopyranoside by means of MS, NMR and nuclear Overhauser effect spectroscopy analyses. A hydrojuglone beta-D-glucopyranoside beta-glucosidase (EC 3.2.1.21) was purified to homogeneity from Juglans regia L. The enzyme catalysed the release of juglone from hydrojuglone beta-D-glucopyranoside with high specificity and showed Michaelis-Menten kinetics with K-m = 0.62 mM and V-max = 14.5 mu kat/mg of protein. This enzyme also showed a higher activity towards beta-D-fucosyl than beta-D-glucosyl bonds. The purified enzyme had an apparent M-r of 64000 by SDS/PAGE and a pI 8.9 by isoelectrofocusing PAGE. The purified enzyme was inhibited by several bivalent cations, such as Cu2+, Fe2+, Hg2+, and by D-glucono-1,5-lactone, showing non-competitive inhibition of the mixed type.