[Evaluation of Conventional Antifungal Agents against Recombinant Yeast Overexpressing β-1,3-Glucanase]

Only 4 classes of antifungal agents comprising 9 compounds are effective against deep mycosis in Japan, and it has been difficult to develop new antifungal specific agents. Micafungin, which has been used as an antifungal agent since 2002, inhibits β-1,3-glucan synthesis in fungal cell walls, thereby killing yeast and filamentous fungi with no septum. In this study, we constructed a pYES2-BGL2 vector to overexpress β-1,3-glucanase (BGL2) in yeast (Saccharomyces cerevisiae INVSc1) and evaluated the synergy between BGL2 overexpression and conventional antifungal agents. The recombinant yeast was incubated in SC-Ura medium, which contained galactose to induce BGL2 overexpression. The recombinant yeast with induced BGL2 overexpression was also frozen and crushed to obtain crude protein for sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), which revealed robust BGL2 overexpression compared with that in the control yeast without the expression vector. Therefore, we considered that we successfully constructed the recombinant yeast to express more BGL2. Further, 3 conventional antifungal agents (amphotericin B, micafungin, and miconazole) were more effective against the recombinant yeast than against the control yeast. From this result, it is suggested that BGL2 overexpression has an enhancing effect on conventional antifungal agents. Hence, glucanase-inducing compounds could act as novel antifungal drugs by augmenting the effectiveness of conventional antifungal agents.