Intensification of Poly(β-L-malic Acid) Production in Aureobasidium melanogenum via ARTP Mutagenesis Through Suppression of Pullulan Biosynthesis

Poly(β-L-malic acid) (PMLA) has attracted considerable industrial attention due to its promising applications in biomedicine, bioplastics, and environmental fields. However, its biosynthesis is highly dependent on elevated dissolved oxygen (DO) levels, while the simultaneous production of pullulan represents a major obstacle. This study introduces a novel strategy to enhance PMLA production in Aureobasidium melanogenum by selectively inhibiting pullulan biosynthesis. We demonstrate that excessive pullulan accumulation severely impairs fermentation performance by significantly reducing oxygen transfer efficiency—an uncharacterized bottleneck in PMLA production. To address this, an ARTP-induced mutant, designated No. H13, was generated, exhibiting an 82.1% reduction in pullulan synthesis. This metabolic shift led to an 86.93% increase in the oxygen mass transfer coefficient (KLa), ultimately enhancing PMLA yield by 72.1% to 45.0 g/L with a specific production of 1.09 g/g. Transcriptomic analysis suggested a potential redirection of carbon flux toward PMLA biosynthesis through coordinated up-regulation of glycolysis and TCA cycle genes, alongside down-regulation of gluconeogenesis and pullulan-exporting ABC transporters. This work presents an alternative to enzymatic approaches by employing a consolidated mutagenesis strategy to reconfigure metabolic networks, offering a strategy for PMLA overproduction.