1. Metabolic engineering of pichia pastoris for malic acid production from methanol.
Wenfang Peng, Min Jiang, Shangjie Zhang, Jiangfeng Ma, Fengxue Xin, Feng Guo, Wenming Zhang, Weiliang Dong, Jie Zhou, Zhongxue Dai. Biotechnol Bioeng. 2021 Jan; 118(1): 357-371. DOI: 10.1002/bit.27575. PMID: 32965690.
The application of rational design in reallocating metabolic flux to accumulate desired chemicals is always restricted by the native regulatory network. In this study, recombinant Pichia pastoris was constructed for malic acid production from sole methanol through rational redistribution of metabolic flux. Different malic acid accumulation modules were systematically evaluated and optimized in P. pastoris. The recombinant PP-CM301 could produce 8.55 g/L malic acid from glucose, which showed a 3.45-fold increase compared to the parent strain. To improve the efficiency of site-directed gene knockout, NHEJ-related protein Ku70 was destroyed, whereas leading to the silencing of heterogenous genes. Hence, genes related to by-product generation were deleted via a specially designed FRT/FLP system, which successfully reduced succinic acid and ethanol production. Furthermore, a key node in the methanol assimilation pathway, glucose-6-phosphate isomerase was knocked out to liberate metabolic fluxes trapped in the XuMP cycle, which finally enabled 2.79 g/L malic acid accumulation from sole methanol feeding with nitrogen source optimization. These results will provide guidance and reference for the metabolic engineering of P. pastoris to produce value-added chemicals from methanol.
2. A study into the role of l-aspartic acid on the metabolism of l-malic acid and d-glucose by oenococcus oeni.
P Jeandet, A Maujean, C Dion, Y Vasserot, E Bonnet. J Appl Microbiol. 2001 Mar; 90(3): 380-7. DOI: 10.1046/j.1365-2672.2001.01255.x. PMID: 11298233.
Aims: The purpose of this work was to study the effect of L-aspartic acid concentration on bacterial growth, D-glucose fermentation and L-malic acid consumption of Oenococcus oeni NCFB 1707. Methods and results: Bacterial cultures were performed in synthetic media. Bacterial growth, D-glucose fermentation and L-malic acid consumption were reduced when L-aspartic acid concentration became excessive. This inhibitory effect of high concentrations of L-aspartic acid on bacterial growth was also observed with several Oenococcus oeni strains, except O. oeni BL01. The L-aspartic acid inhibitory effect on bacterial growth could be reduced by increasing the concentration of L-glutamic acid. L-glutamic acid transport was found to be competitively inhibited by L-aspartic acid. In addition, an excessive amount of L-aspartic acid modified D-glucose metabolism, with an overproduction of acetic acid and reduced ethanol production. Conclusion: Since L-glutamic acid is an essential amino acid for the bacterial strain used, the L-aspartic acid inhibitory effect on bacterial growth could be linked to its involvement in an antagonistic interaction with L-glutamic acid. Significance and impact of the study: Such antagonistic interactions between amino acids in O. oeni strains could be another explanation for the difficulties of inducing malolactic fermentation in wines.
3. Efficacy of malic acid against listeria monocytogenes attached to poultry skin during refrigerated storage.
B Herrera, E González-Fandos. Poult Sci. 2013 Jul; 92(7): 1936-41. DOI: 10.3382/ps.2012-02968. PMID: 23776283.
This work evaluated the effect of malic acid washing on the growth of Listeria monocytogenes on poultry legs stored at 4°C for 8 d. Fresh inoculated chicken legs were dipped into a 1 or 2% malic acid solution (vol/vol) for 5 min or distilled water (control). Surface pH values, sensorial characteristics (odor, color, texture, and overall appearance) and L. monocytogenes, mesophile, psychrotroph, and Enterobacteriaceae counts were evaluated after treatment (d 0) and after 1, 3, 6, and 8 d of storage at 4°C. Legs washed with 2% malic acid showed a significant (P < 0.05) inhibitory effect on L. monocytogenes compared with control legs, with a decrease of about 1.66 log units after treatment. Sensory quality was not adversely affected by malic acid. Treatments with malic acid reduced bacterial growth and preserved reasonable sensorial quality after storage at 4°C for 6 d. This study demonstrates that, although malic acid did reduce populations of L. monocytogenes on poultry, it did not completely inactivate the pathogen. The application of malic acid may be used as an additional hurdle contributing to extend the shelf life of raw poultry.
