NADPH-dependent 5-keto-D-gluconate reductase is a part of the fungal pathway for D-glucuronate catabolism

Abstract

NADPH-dependent 5-keto-D-gluconate reductase was identified as a missing element in the pathway for D-glucuronate catabolism in fungi. The disruption of the gene, gluF, by CRISPR/Cas9 in the filamentous fungus Aspergillus niger resulted in a strain unable to catabolise D-glucuronate. The purified GluF protein was characterized and k_cat and K_m values of 23.7 ± 1.8 s^-1 and 3.2 ± 0.1 mm for 5-keto-D-gluconate, respectively, were determined. The enzyme is reversible and is active with NADP⁺ and D-gluconate. We suggest a pathway for D-glucuronate catabolism with the intermediates L-gulonate, 2-keto-L-gulonate, L-idonate, 5-keto-D-gluconate, D-gluconate and D-gluconate-6-phosphate which is a part of the pentose phosphate pathway. A fungal enzyme activity for the conversion of L-gulonate to 2-keto-L-gulonate remains to be identified.

Keywords: Aspergillus niger; 5-keto-gluconate; CRISPR/Cas9; D-gluconate-5-dehydrogenase; D-glucuronate; EC 1.1.1.69; fungal pathway.