Analysis of the L-malate biosynthesis pathway involved in poly(β-L-malic acid) production in Aureobasidium melanogenum GXZ-6 by addition of metabolic intermediates and inhibitors.

Abstract

Poly(β-L-malic acid) (PMA) is a promising polyester formed from L-malate in microbial cells. Malate biosynthesis is crucial for PMA production. Previous studies have shown that the non-oxidative pathway or oxidative pathway (TCA cycle) is the main biosynthetic pathway of malate in most of PMA-producing strains, while the glyoxylate cycle is only a supplementary pathway. In this study, we investigated the effect of exogenous metabolic intermediates and inhibitors of the malate biosynthetic pathway on PMA production by Aureobasidium melanogenum GXZ-6. The results showed that PMA production was stimulated by maleic acid (a fumarase inhibitor) and sodium malonate (a succinate dehydrogenase inhibitor) but inhibited by succinic acid and fumaric acid. This indicated that the TCA cycle might not be the only biosynthetic pathway of malate. In addition, the PMA titer increased by 18.1% upon the addition of glyoxylic acid after 72 h of fermentation, but the PMA titer decreased by 7.5% when itaconic acid (an isocitrate lyase inhibitor) was added, which indicated that malate for PMA production was synthesized significantly via the glyoxylate cycle rather than the TCA cycle. Furthermore, in vitro enzyme activities of the TCA and glyoxylate cycles suggested that the glyoxylate cycle significantly contributed to the PMA production, which is contradictory to what has been reported previously in other PMA-producing A. pullulans.