Corynebacterium glutamicum as platform for the production of hydroxybenzoic acids

Abstract

BackgroundHydroxybenzoic acids are industrially relevant aromatic compounds, which also play key roles in the microbial carbon metabolism, e.g., as precursors for the synthesis of cofactors or metal-chelating molecules. Due to its pronounced resistance to aromatics Corynebacterium glutamicum represents an interesting platform for production of these compounds. Unfortunately, a complex catabolic network for aromatic molecules prevents application of C. glutamicum for microbial production of aromatic compounds other than aromatic amino acids, which cannot be metabolized by this microorganism.ResultsWe completed the construction of the platform strain C. glutamicum DelAro5, in which the deletion of altogether 27 genes in five gene clusters abolished most of the peripheral and central catabolic pathways for aromatic compounds known in this microorganism. The obtained strain was subsequently applied for the production of 2-hydroxybenzoate (salicylate), 3-hydroxybenzoate, 4-hydroxybenzoate and protocatechuate, which all derive from intermediates of the aromatic amino acid-forming shikimate pathway. For an optimal connection of the designed hydroxybenzoate production pathways to the host metabolism, C. glutamicum was additionally engineered towards increased supply of the shikimate pathway substrates erythrose-4-phosphate and phosphoenolpyruvate by manipulation of the glucose transport and key enzymatic activities of the central carbon metabolism. With an optimized genetic background the constructed strains produced 0.01 g/L (0.07 mM) 2-hydroxybenzoate, 0.3 g/L (2.2 mM) 3-hydroxybenzoate, 2.0 g/L (13.0 mM) protocatechuate and 3.3 g/L (23.9 mM) 4-hydroxybenzoate in shaking flasks.ConclusionBy abolishing its natural catabolic network for aromatic compounds, C. glutamicum was turned into a versatile microbial platform for aromatics production, which could be exemplarily demonstrated by rapidly engineering this platform organism towards producing four biotechnologically interesting hydroxybenzoates. Production of these compounds was optimized following different metabolic engineering strategies leading to increased precursor availability. The constructed C. glutamicum strains are promising hosts for the production of hydroxybenzoates and other aromatic compounds at larger scales.Electronic supplementary materialThe online version of this article (10.1186/s12934-018-0923-x) contains supplementary material, which is available to authorized users.

Metabolic and evolutionary responses of Clostridium thermocellum to genetic interventions aimed at improving ethanol production

“…Lee Wen and Bao [57] used pathway engineering targeting metabolic control to produce 62.5 g/L lysine from corn stover hydrolysate. C. glutamicum has been developed to the point that it may plausibly be considered as an alternative to E. coli as a platform for products other than amino acids [58,59].…”

Evaluation of process involved in the production of aromatic compounds in Gram‐negative bacteria isolated from vanilla ( Vanilla planifolia ex. Andrews) beans

“…Moreover some micro‐organisms can produce or degrade AC like vanillin and HBA. Also, there are Gram‐negative and Gram‐positive bacteria that produce vanillin from FA like precursors and, on rare occasions, they produce HBA and can utilize AC as a defense mechanism to inhibit competing bacterial growth (Chen et al ; Kallscheuer and Marienhagen ; Sannino et al ; Shin et al ).…”

Competence ofCorynebacterium glutamicumas a host for the production of type I polyketides

“…glutamicum and demonstrated its suitability as a cell factory for 6-MSA synthesis. Production of related hydroxybenzoic acids in engineered microorganism such as salicylic acid (a derivative of 6-MSA lacking the methyl group) requires deregulation and engineering of the shikimate pathway and can lead to metabolic imbalances, auxotrophic strains or undesired accumulation of side products (Kallscheuer and Marienhagen, 2018;Lin et al, 2014). In contrast, after introduction of ChlB1 Sa in C. glutamicum no significant accumulation of side products could be observed.…”

“…Since decades, Corynebacterium glutamicum is used at industrial scale for the production of amino acids, in particular of L-glutamate and L-lysine (Eggeling and Bott, 2015), but this bacterium was also engineered for the synthesis of other biotechnologically interesting compounds, e.g. alcohols, diamines, dicarboxylic acids, aromatic compounds and secondary metabolites (Heider et al, 2014;Kallscheuer and Marienhagen, 2018;Kallscheuer et al, 2016a;Litsanov et al, 2012;Nguyen et al, 2015;Vogt et al, 2016). In previous studies, we already established the functional introduction of plant-derived type III PKSs with a typically length of 300-400 amino acids into C. glutamicum, which enabled plant polyphenol synthesis with this bacterium (Kallscheuer et al, 2016b;Milke et al, 2019b).…”