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Literature summary extracted from

  • Dahal, G.P.; Viola, R.E.
    Structural insights into inhibitor binding to a fungal ortholog of aspartate semialdehyde dehydrogenase (2018), Biochem. Biophys. Res. Commun., 503, 2848-2854 .
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.2.1.11 gene BDCG_01946, sequence comparisons, recombinant expression of His-tagged enzyme in Escherichia coli Blastomyces dermatitidis

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
1.2.1.11 purified recombinant His6-tagged enzyme free or in complex with NADP+ and inhibitor 4-benzoquinone, hanging drop vapor diffusion, mixing of 12 mg/ml protein solution with reservoir solution containing 0.2 M ammonium citrate, pH 7.0, and 18% PEG 3350, for inhibitor complexed crystals soaking in reservoir solution with added 15% v/v ethylene glycol and 5 mM NADP+, X-ray diffraction structure determination and analysis at 2.4-2.6 A resolution, molecular replacement using the structure of Candida albicans ASADH (PDB ID 3hsk) as the search model Blastomyces dermatitidis

General Stability

EC Number General Stability Organism
1.2.1.11 NADP+ binding stabilizes the dimer of dimers necessary for catalytic activity in fungal ASADHs Blastomyces dermatitidis

Inhibitors

EC Number Inhibitors Comment Organism Structure
1.2.1.11 2-chloro-3-methoxy-1,4-naphthoquinone a selective, low to sub-micromolar inhibitors of the fungal ASADHs Blastomyces dermatitidis
1.2.1.11 4-benzoquinone enzyme binding structure analysis, a competitive and rel. weak inhibitor. The carbonyl oxygens of the inhibitor each participate in hydrogen bonds, one with the epsilon-nitrogen of Lys211 and the guanidine nitrogen of Arg114 (substrate binding residues), and the other with the thiol of the Cys154 nucleophile. There are also hydrophobic interactions with Asn153. While the position of these carbonyl groups are fixed, the aromatic ring adopts a somewhat different orientation in each of the subunits of the dimer Blastomyces dermatitidis

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.2.1.11 L-aspartate 4-semialdehyde + phosphate + NADP+ Blastomyces dermatitidis
-
 L-4-aspartyl phosphate + NADPH + H+
-
 r
1.2.1.11 L-aspartate 4-semialdehyde + phosphate + NADP+ Blastomyces dermatitidis ER-3
-
 L-4-aspartyl phosphate + NADPH + H+
-
 r
1.2.1.11 L-aspartate 4-semialdehyde + phosphate + NADP+ Blastomyces dermatitidis ATCC MYA-2586
-
 L-4-aspartyl phosphate + NADPH + H+
-
 r

Organism

EC Number Organism UniProt Comment Textmining
1.2.1.11 Blastomyces dermatitidis C5GC63
-
-
1.2.1.11 Blastomyces dermatitidis ATCC MYA-2586 C5GC63
-
-
1.2.1.11 Blastomyces dermatitidis ER-3 C5GC63
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.2.1.11 recombinant His-tagged enzyme from Escherichia coli by nickel affinity and anion exchange chromatography, followed by ultrafiltration Blastomyces dermatitidis

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.2.1.11 L-aspartate 4-semialdehyde + phosphate + NADP+
-
 Blastomyces dermatitidis L-4-aspartyl phosphate + NADPH + H+
-
 r
1.2.1.11 L-aspartate 4-semialdehyde + phosphate + NADP+
-
 Blastomyces dermatitidis ER-3 L-4-aspartyl phosphate + NADPH + H+
-
 r
1.2.1.11 L-aspartate 4-semialdehyde + phosphate + NADP+
-
 Blastomyces dermatitidis ATCC MYA-2586 L-4-aspartyl phosphate + NADPH + H+
-
 r

Subunits

EC Number Subunits Comment Organism
1.2.1.11 homotetramer dimer-of-dimers Blastomyces dermatitidis

Synonyms

EC Number Synonyms Comment Organism
1.2.1.11 ASADH
-
 Blastomyces dermatitidis
1.2.1.11 aspartate semialdehyde dehydrogenase
-
 Blastomyces dermatitidis
1.2.1.11 BDCG_01946
-
 Blastomyces dermatitidis

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.2.1.11 25
-
 assay at Blastomyces dermatitidis

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.2.1.11 8.6
-
 assay at Blastomyces dermatitidis

Cofactor

EC Number Cofactor Comment Organism Structure
1.2.1.11 NADP+ NADP+ binding stabilizes the dimer of dimers necessary for catalytic activity in fungal ASADHs Blastomyces dermatitidis
1.2.1.11 NADPH the nicotinamide moiety near the active site cysteine also places it in an optimal position for hydride transfer from NADPH to the enzyme-bound intermediate during the catalytic cycle Blastomyces dermatitidis

Ki Value [mM]

EC Number Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
1.2.1.11 0.126
-
 4-benzoquinone pH 8.6, 25°C, recombinant enzyme Blastomyces dermatitidis

General Information

EC Number General Information Comment Organism
1.2.1.11 additional information the nicotinamide moiety near the active site cysteine also places it in an optimal position for hydride transfer from NADPH to the enzyme-bound intermediate during the catalytic cycle Blastomyces dermatitidis