Aokap9 gene knockout contributes to kojic acid synthesis in Aspergillus oryzae.

Background: Kojic acid, a significant secondary metabolite primarily synthesized by Aspergillus oryzae, has extensive applications across various industries. It is imperative to identify effective gene targets that can enhance kojic acid production and to elucidate its biosynthetic regulation for yield optimization in A. oryzae.

Results: In this study, we identify the Aokap9 gene, which encodes an uncharacterized protein containing two transmembrane domains, as being closely associated with kojic acid synthesis. Disruption of Aokap9 resulted in increased kojic acid production, whereas mutations in either kojR or laeA within the ΔAokap9 background abolished synthesis, indicating that Aokap9 regulates kojic acid synthesis through kojR and laeA. Conversely, AozfA overexpression suppressed kojic acid accumulation in the ΔAokap9 strain. Furthermore, disruption of AozfA or overexpression of kojR in the Aokap9 mutant resulted in significantly elevated yields, unlike the effects observed with laeA overexpression. Transcriptome profiling further revealed that AoKap9 plays a role in the oxidative stress response through influencing the expression of superoxide dismutase, catalase, and copper amine oxidase in A. oryzae.

Conclusion: AoKap9 mediates kojic acid synthesis through a regulatory pathway involving AoZFA, LaeA, and KojR, making it as an important gene target for enhancing kojic acid in A. oryzae. These findings provide essential insights into the regulatory mechanisms governing kojic acid biosynthesis and highlight Aokap9 as a promising target for metabolic engineering in A. oryzae.