Improved Production of Malic Acid in Aspergillus niger by Abolishing Citric Acid Accumulation and Enhancing Glycolytic Flux.

Improved Production of Malic Acid in Aspergillus niger by Abolishing Citric Acid Accumulation and Enhancing Glycolytic Flux.

ACS Synth Biol. 2020 May 07;:

Authors: Xu Y, Zhou Y, Cao W, Liu H

Abstract
Microbial fermentation was widely explored to produce malic acid. Previously, Aspergillus niger has been successfully engineered and high titer of malic acid was achieved with strain S575, but it also produced a high level of by-product citric acid. Here, the capability of A. niger in malic acid biosynthesis was further improved by eliminating the accumulation of citric acid and enhancing glycolytic flux. Characterization of variant mutants suggested that disruption of cexA, a gene encoding citric acid transporter located on cell membrane, abolished citric acid accumulation. However, cexA-deficient strain S895 showed significantly decreased malic acid production. Further analysis of S895 indicated that the transcription level of genes involved in glucose transportation and glycolytic pathway was significantly reduced and the corresponding enzyme activity was also lower than those of S575. Individual overexpression of genes encoding glucose transporter MstC and key enzymes (hexokinase HxkA, 6-phosphofructo-2-kinase PfkA and pyruvate kinase PkiA) involved in irreversible reactions of glycolic pathway increased malic acid production. Accordingly, genes of mstC, hxkA, pfkA and pkiA were overexpressed altogether in S895 and the resultant strain S1149 was constructed. The titer of malic acid in batch fermentation with S1149 reached 201.13 g/L. Compared with S575, the by-product of citric acid was completely abolished in S1149, and the ratio of malic acid/glucose was increased from 1.27 to 1.64 mol/mol, the highest yield reported as so far and fermentation period was shorted from 9 d to 8 d. Thus, a strain with great industrial application potential was developed by engineering 9 genes in A. niger and a pilot fermentation technology was exploited.

PMID: 32379964 [PubMed – as supplied by publisher]

Source: Industry