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Literature summary for 1.1.1.365 extracted from

  • Harth, S.; Wagner, J.; Sens, T.; Choe, J.Y.; Benz, J.P.; Weuster-Botz, D.; Oreb, M.
    Engineering cofactor supply and NADH-dependent D-galacturonic acid reductases for redox-balanced production of L-galactonate in Saccharomyces cerevisiae (2020), Sci. Rep., 10, 19021 .
    View publication on PubMedView publication on EuropePMC
Search for more literature for 1.1.1.365

Application

Application Comment Organism
synthesis L-galactonate shows potential applications as an additive to nutrients and cosmetics. AnGar1 variants are developed that represent the a GalA reductases with a higher preference for NADH compared to NADPH. The altered cofactor-specificity enables the coupling of GalA reduction to glycolysis, resulting in higher yields of GalOA when glucose is used as a redox donor. The engineered AnGar1 should prove valuable for D-galacturonic acid utilization in pectin-rich hydrolysates, which contain neutral sugars such as glucose, galactose, or arabinose, all of which are funneled into glycolysis. The NADH-dependent GalA reductases can facilitate the coupling of L-galactonate production to the oxidation of glycerol, an abundant waste product that can be supplemented to pectin-rich hydrolysates Aspergillus niger

Cloned(Commentary)

Cloned (Comment) Organism
expression in Saccharomyces cerevisie Aspergillus niger
gene gaaA, recombinant expression of wild-type enzyme in Saccharomyces cerevisiae strain SiHY001 Aspergillus niger
gene gar1, recombinant expression of wild-type and mutant enzymes in Saccharomyces cerevisiae strain SiHY001, real-time PCR expression analysis Aspergillus niger

Protein Variants

Protein Variants Comment Organism
K261M site-directed mutagenesis, the recombinant yeast strain expressing the enzyme mutant shows increased activity with NADH in L-galacturonate reduction compared to wild-type Aspergillus niger
K261M the mutation confers AnGar1 the ability to accept NADH in addition to NADPH. Mutation partly abolishes the AnGar1 activity with NADPH as the cofactor Aspergillus niger
K261M/R267L mutation confers NADH specificity to the enzyme. Mutation almost fully abolishes the AnGar1 activity with NADPH as the cofactor Aspergillus niger
K261M/R267L site-directed mutagenesis, the recombinant yeast strain expressing the enzyme mutant shows increased activity with NADH in L-galacturonate reduction compared to wild-type Aspergillus niger
additional information establishment of the production of L-galactonate (GalOA) or the full GalA catabolic pathway in Saccharomyces cerevisiae using the enzyme mutant K261M/R267L with increased NADH activity, and coupling the reduction of GalA to the oxidation of the sugar alcohol sorbitol that has a higher reduction state compared to glucose for yielding the necessary redox cofactors. By choosing a suitable sorbitol dehydrogenase, yeast strains are designed in which the sorbitol metabolism yields a surplus of either NADPH or NADH Aspergillus niger
R267L site-directed mutagenesis, the recombinant yeast strain expressing the enzyme mutant shows increased activity with NADH in L-galacturonate reduction compared to wild-type Aspergillus niger
R267L the mutation confers AnGar1 the ability to accept NADH in addition to NADPH. Mutation partly abolishes the AnGar1 activity with NADPH as the cofactor Aspergillus niger

Inhibitors

Inhibitors Comment Organism Structure
NADP+ mutant enzyme R267L shows a considerable sensitivity at the lower (0.16 mM) but not at the higher (0.8 mM) NADH concentration Aspergillus niger

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
D-galacturonate + NADPH + H+ Aspergillus niger
-
 L-galactonate + NADP+
-
 ?
D-galacturonate + NADPH + H+ Aspergillus niger CBS 513.88
-
 L-galactonate + NADP+
-
 ?
L-galactonate + NAD+ Aspergillus niger
-
 D-galacturonate + NADH + H+
-
 r
L-galactonate + NADP+ Aspergillus niger
-
 D-galacturonate + NADPH + H+
-
 r

Organism

Organism UniProt Comment Textmining
Aspergillus niger
-
-
-
Aspergillus niger A2R7U3
-
-
Aspergillus niger A8DRH9
-
-
Aspergillus niger CBS 513.88 A2R7U3
-
-

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
D-galacturonate + NADPH + H+
-
 Aspergillus niger L-galactonate + NADP+
-
 ?
D-galacturonate + NADPH + H+
-
 Aspergillus niger CBS 513.88 L-galactonate + NADP+
-
 ?
L-galactonate + NAD+
-
 Aspergillus niger D-galacturonate + NADH + H+
-
 r
L-galactonate + NADP+
-
 Aspergillus niger D-galacturonate + NADPH + H+
-
 r

Synonyms

Synonyms Comment Organism
AnGaaA
-
 Aspergillus niger
AnGar1
-
 Aspergillus niger
D-galacturonic acid reductase
-
 Aspergillus niger
D-galacturonic acid reductases
-
 Aspergillus niger
gaaA
-
 Aspergillus niger
GalA reductase
-
 Aspergillus niger
gar1
-
 Aspergillus niger

Cofactor

Cofactor Comment Organism Structure
additional information the enzyme accepts NADPH as well as NADH as cofactor. But its catalytic efficiency for GalA reduction is approximately 50fold higher with NADPH than with NADH Aspergillus niger
additional information the wild-type enzyme Gar1 accepts NADPH but not NADH as cofactor Aspergillus niger
NAD+
-
 Aspergillus niger
NADH
-
 Aspergillus niger
NADP+
-
 Aspergillus niger
NADPH
-
 Aspergillus niger

General Information

General Information Comment Organism
evolution AnGaaA is identified as the bona fide GalA reductase in Aspergillus niger. AnGaaA is not related to Gar1 proteins Aspergillus niger