The pathway intermediate 2‐keto‐3‐deoxy‐L‐galactonate mediates the induction of genes involved in D‐galacturonic acid utilization in Aspergillus niger

Abstract

In Aspergillus niger, the enzymes encoded by gaaA, gaaB, and gaaC catabolize d‐galacturonic acid (GA) consecutively into l‐galactonate, 2‐keto‐3‐deoxy‐l‐galactonate, pyruvate, and l‐glyceraldehyde, while GaaD converts l‐glyceraldehyde to glycerol. Deletion of gaaB or gaaC results in severely impaired growth on GA and accumulation of l‐galactonate and 2‐keto‐3‐deoxy‐l‐galactonate, respectively. Expression levels of GA‐responsive genes are specifically elevated in the ∆gaaC mutant on GA as compared to the reference strain and other GA catabolic pathway deletion mutants. This indicates that 2‐keto‐3‐deoxy‐l‐galactonate is the inducer of genes required for GA utilization.

W361R mutation in GaaR, the regulator of D‐galacturonic acid‐responsive genes, leads to constitutive production of pectinases in Aspergillus niger

“…The subcellular localization of the C‐terminally eGFP‐tagged GaaR and GaaR W361R in gaaR‐eGFP (EA36.1) and gaaR W361R ‐eGFP (EA39.1) grown under inducing (i.e., on 2‐keto‐3‐deoxy‐L‐galactonate) or non‐inducing (i.e., on D‐fructose) conditions was analyzed using confocal laser scanning microscopy (Figure ). Beforehand, the potency of 2‐keto‐3‐deoxy‐L‐galactonate, the physiological inducer (Alazi et al., ), to induce the expression of the GA‐responsive genes was tested using the promoter–reporter strain P pgaX ‐ amdS ∆creA (JN29.2) and was compared to that of GA (Figure S5). Addition of 100 nM 2‐keto‐3‐deoxy‐L‐galactonate to the growth media resulted in a major increase in growth, whereas addition of a 100 nM GA was not sufficient to induce the growth of P pgaX ‐ amdS ∆creA (JN29.2), indicating that 2‐keto‐3‐deoxy‐L‐galactonate is able to induce pgaX expression and that 2‐keto‐3‐deoxy‐L‐galactonate is more potent than GA in inducing the expression.…”

“…Hawkins, Lamb, Moore, Charles, and Roberts () have proposed that the QutR domain homologous to the AROM 3‐dehydroquinate dehydratase can recognize and bind to dehydroquinic acid produced from quinic acid, but does not react on it enzymatically in A. nidulans . For A. niger , accumulation of 2‐keto‐3‐deoxy‐L‐galactonate in Δ gaaC results in hyper‐induction of GA‐responsive genes and the inducer 2‐keto‐3‐deoxy‐L‐galactonate is likely to interact with the GaaX repressor protein (Alazi et al., ). The GA catabolic pathway enzyme GaaC is an aldolase and converts 2‐keto‐3‐deoxy‐L‐galactonate to pyruvate and L‐glyceraldehyde.…”

“…Constitutive production of plant biomass degrading enzymes like pectinases is interesting for the industrial production of these enzymes because it allows the production independently of the feedstock. By understanding the transcriptional regulatory mechanism of pectinases, several approaches have been conducted leading to inducer independent and abundant production of pectinases, such as deletion of GaaX (Niu et al., ), overexpression of GaaR from a constitutive promoter (Alazi et al., ), or accumulation of the inducer (Alazi et al., ). Here, we show that a mutation in GaaR can also lead to constitutive production of pectinases.…”

“…This suggests that the inducer binds to GaaX resulting in a GaaX‐unbound and active form of GaaR under inducing conditions. The catabolic intermediate 2‐keto‐3‐deoxy‐L‐galactonate produced from GA was shown to induce the production of pectinases by A. niger which makes it reasonable to suggest that 2‐keto‐3‐deoxy‐L‐galactonate binds to GaaX resulting in the dissociation of GaaX from GaaR, thereby releasing GaaR as an active transcription factor (Alazi et al., ). Because overexpression of the activators involved in GA or quinic acid utilization has similar effects, it is likely that activation mechanism of both QutA/QutR and GaaR/GaaX is conserved and comparable.…”

Basidiomycotic Yeast Cryptococcus diffluens Converts l-Galactonic Acid to the Compound on the Similar Metabolic Pathway in Ascomycetes

“…However, it is not known what is produced following l-GalA conversion in the d-GalUA metabolic pathway of C. diffluens. It has been shown in ascomycetes that one metabolic pathway from l-GalA processes a dehydratase reaction that converts l-GalA to l-threo-3-deoxy-hexulosonic acid/2-keto-3-deoxy-l-galactonic acid (l-2KDGal) and then, an aldolase reaction converts l-2KDGal to l-glycerol and pyruvic acid [8][9][10]. Another pathway from l-GalA results in the conversion to l-ascorbic acid, which has been shown in Cryptococcus laurentii, which is closely related to C. diffluens [11].…”