Characterizationof Myrosinase-Mediated GlucosinolateDegradation Pathways in <i>Lactiplantibacillus plantarum</i> ZUST49

Cruciferous vegetables are rich in glucosinolates that can be hydrolyzed by myrosinase into isothiocyanates (ITCs) with significant anticancer properties. In the absence of bacterial myrosinase, glucosinolates are excreted from the body in their inactive forms. However, the mechanisms underlying the bacterial breakdown of glucosinolates are not well understood. Here, we investigated the mechanism and enzymes involved in glucosinolate breakdown by the probiotic microorganism <i>Lactiplantibacillus plantarum</i> ZUST49, which degrades the glucosinolate glucoraphanin to sulforaphane and erucin. The glucoraphanin-degrading activity of this strain was induced by the presence of glucoraphanin and an absence of glucose. UPLC-MS analysis of the degradation products indicated that glucoraphanin was degraded via three distinct pathways, and further, transcriptomic and proteomic analyses led to the identification of a myrosinase gene, <i>LpMyr</i>, that encodes a 460-amino acid enzyme. The purified <i>Lp</i>Myr protein exhibited optimal activity at 50 °C and pH 7.0, with hydrolysis rates of 7.74 U/mg for glucoraphanin and 5.89 U/mg for sinigrin. These findings provide new insights into the glucosinolate conversion capability of <i>L. plantarum</i> and highlight its potential for high-yield ITC production in the fermentation industry, as well as its potential use as a probiotic in the human gut.