Extracellular enzymes secreted in the mycelial block of Lentinula edodes during hyphal growth

Lentinula edodes (shiitake mushroom) is one of the most widely cultivated edible mushrooms and is primarily cultivated using sawdust medium. While there have been improvements in the cultivation technology, the mechanism of mycelial block cultivation, such as mycelial growth and enzymatic sawdust degradation, has not been clarified. In this study, the mycelium was elongated longitudinally in the bottle sawdust culture for 27 days, and the cultivated sawdust medium was divided into three sections (top, middle, and bottom parts). To determine spatial heterogeneity in the enzyme secretion, the enzymatic activities of each part were analyzed. Lignocellulose degradation enzymes, such as endoglucanase, xylanase, and manganese peroxidase were highly secreted in the top part of the medium. On the other hand, amylase, pectinase, fungal cell wall degradation enzyme (β-1,3-glucanase, β-1,6-glucanase, and chitinase), and laccase activities were higher in the bottom part. The results indicate that the principal sawdust degradation occurs after mycelial colonization. Proteins with the laccase activity were purified from the bottom part of the medium, and three laccases, Lcc5, Lcc6 and Lcc13, were identified. In particular, the expression of Lcc13 gene was higher in the bottom part compared with the level in the top part, suggesting Lcc13 is mainly produced from the tip region and have important roles for mycelial spread and nutrient uptake during early stage of cultivation.