Comparative elucidation of properties of sucrase-cellobiase co-aggregate produced in media containing sucrose by <i style="">Termitomyces clypeatus</i>

Abstract

Production profiles and characterization of the sucrase-cellobiase (S-C) co-aggregates from the filamentous fungus <i style="">Termitomyces clypeatus</i> were compared in media containing 1% and 5% sucrose to understand the effect of cellular regulation over secretion and aggregation of these two industrially important glycosidases. The enzymes were secreted constitutively but in a high sucrose medium (5%), cellobiase secretion was reduced to a basal level of 0.03 U/mL. In intracellular, cell-bound and extracellular milieus, S/C ratios gradually declined in a predictable trend indicating participation of more cellobiase subunits for secretion. Sucrase was secreted <i style="">via</i> vacuoles in the fungus, following the same route as that of cellobiase and thus co-aggregates of S-C were present in the vacuolar fraction. The extracellular co-aggregates showed similar molecular sizes (>550 kDa) on zymography; however, SDS-PAGE revealed substantial difference in their subunit assemblies. Sucrase from the 5% medium showed a 2.6 times lower K_m than 1% medium. These observations demonstrated the formation of a unique S-C co-assembly, optimally suited to its needs and accommodation of the constituent subunits, to be used for biotechnological applications.