Enzymic properties of purified myrosinase from lepidium sativum seedlings
To establish the substrate specificity of the thioglucoside glucohydrolase myrosinase (EC 184.108.40.206), this enzyme was purified to homogeneity from light-grown cress (Lepidium sativum L.) seedlings by Sephadex gel filtration. Red Dye and anion exchange (FPLC Mono Q) chromatography, and preparative isoelectric focusing. Hydrolytic activity was shown toward only 4 of the 29 synthetic and natural O and S-glycosides tested. Highest activity was displayed with the endogenous glucosinolates benzylglucosinolate (Km, 295 μM) and sinigrin (Km, 300 μM) at an optimum pH of 5.5 in sodium citrate buffer. The synthetic glycosides PNPG (Km, 2.0 μM) and ONPG were poorer substrates at an optimum pH of 6.5 in potassium phosphate buffer. The enzyme was inactive with all other nitrophenyl glycosides tested including PNP-α-D-glucoside and PNP-thio-β-D-glucoside, suggesting a requirement for O-β-D-glucose as the glycone moiety within these substrates. PNPG hydrolysis was stimulated 2.6-fold by ascorbate (1 mM). The enzyme exhibited no metal ion requirement and was strongly inhibited by lead nitrate, mercury chloride, and ferric chloride at 1 mM concentration. The metal chelators DIECA, EDTA, o-phenanthroline, and 2, 2′-dipyridyl were not inhibitory, but the thiol reagents PCM S, PCMB, and N-ethylmaleimide (at 1 mM) caused 50-80% inhibition of enzyme activity.
Glucosinolate Degradation, Lepidium sativum, Myrosinase, Substrate Specificity, Thioglucoside Glucohydrolase
Durham, Paul L., and Jonathan E. Poulton. "Enzymic properties of purified myrosinase from Lepidium sativum seedlings." Zeitschrift für Naturforschung C 45, no. 3-4 (1990): 173-178.
Zeitschrift fur Naturforschung - Section C Journal of Biosciences