Baicalein Acts against Candida albicans by Targeting Eno1 and Inhibiting Glycolysis

Baicalein (BE) is a promising antifungal small-molecule compound with an extended antifungal spectrum, good synergy with fluconazole, and low toxicity, but its target protein and antifungal mechanism remain elusive. In this study, we found that BE can function against Candida albicans by disrupting glycolysis through targeting Eno1 and inhibiting its function. Eno1 acts as a key therapeutic target of the drug, as BE had no antifungal activity against the eno1 null mutant in a Galleria mellonella model of C. albicans infection. To investigate the mechanism of action, we solved the crystal structure of C. albicans Eno1(CaEno1) and then compared the difference between this structure and that of Eno1 from humans. The predicted primary binding site of BE on CaEno1 is between amino acids D261 and W274, with D263, S269, and K273 playing critical roles in the interaction with BE. Both positions S269 and K273 have different residues in the human Eno1 (hEno1). This finding suggests that BE may bind selectively to CaEno1, which would limit the potential for side effects in humans. Our findings demonstrate that Eno1 is a target protein of BE and thus may serve as a novel target for the development of antifungal therapeutics acting through the inhibition of glycolysis. IMPORTANCE Baicalein (BE) is a promising antifungal agent which has been well characterized, but its target protein is still undiscovered. The protein Eno1 plays a crucial role in the survival of Candida albicans. However, there are few antifungal agents which inhibit the functions of Eno1. Here, we found that BE can function against Candida albicans by disrupting glycolysis through targeting Eno1 and inhibiting its function. We further solved the crystal structure of C. albicans Eno1(CaEno1) and predicted that the primary binding site of BE on CaEno1 is between amino acids D261 and W274, with D263, S269, and K273 playing critical roles in the interaction with BE. Our findings will be helpful to get specific small-molecule inhibitors of CaEno1 and open the way for the development of new antifungal therapeutics targeted at inhibiting glycolysis.