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Literature summary extracted from

  • Volodina, E.; Steinbuechel, A.
    (S)-3-hydroxyacyl-CoA dehydrogenase/enoyl-CoA hydratase (FadB) from fatty acid degradation operon of Ralstonia eutropha H16 (2014), AMB Express, 4, 69.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.1.1.35 DNA and amino acid sequence determination and analysis, genetic organization, sequence comparisons and phylogenetic tree of 3-hydroxyacyl-CoA dehydrogenases, recombinant expression of N-terminally His-tagged enzyme in Escherichia coli strain BL21(DE3) Cupriavidus necator

Inhibitors

EC Number Inhibitors Comment Organism Structure
1.1.1.35 acetyl-CoA inhibits the enzyme by 50% at 0.1 mM, further increase of inhibitor concentration reduces the activity of FadB' drastically Cupriavidus necator
1.1.1.35 CoA inhibits the enzyme by 20% at 0.1 mM, further increase of inhibitor concentration reduces the activity of FadB' drastically Cupriavidus necator
1.1.1.35 propionyl-CoA
-
 Cupriavidus necator

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.1.1.35 0.048
-
 acetoacetyl-CoA pH 6.5, 30°C, recombinant enzyme Cupriavidus necator
1.1.1.35 0.076
-
 acetoacetyl-CoA pH 6.5, 30°C, recombinant enzyme in presence of 0.1 mM acetyl-CoA Cupriavidus necator

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
1.1.1.35 85900
-
-
 Cupriavidus necator
1.1.1.35 86000
-
-
 Cupriavidus necator
1.1.1.35 87000
-
 recombinant His-tagged enzyme, gel filtration Cupriavidus necator

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.1.1.35 (S)-3-hydroxybutanoyl-CoA + NAD+ Cupriavidus necator
-
 acetoacetyl-CoA + NADH + H+
-
 r
1.1.1.35 (S)-3-hydroxybutanoyl-CoA + NAD+ Cupriavidus necator DSM 428
-
 acetoacetyl-CoA + NADH + H+
-
 r

Organism

EC Number Organism UniProt Comment Textmining
1.1.1.35 Cupriavidus necator Q0KEG0
-
-
1.1.1.35 Cupriavidus necator DSM 428 Q0KEG0
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.1.1.35 recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography Cupriavidus necator

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
1.1.1.35 1.5
-
 purified recombinant enzyme, pH 6.5, 30°C, substrate (S)-3-hydroxybutanoyl-CoA with NADP+ Cupriavidus necator
1.1.1.35 16.5
-
 purified recombinant enzyme, pH 6.5, 30°C, substrate (S)-3-hydroxybutanoyl-CoA with NAD+ Cupriavidus necator

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.1.1.35 (S)-3-hydroxybutanoyl-CoA + NAD+
-
 Cupriavidus necator acetoacetyl-CoA + NADH + H+
-
 r
1.1.1.35 (S)-3-hydroxybutanoyl-CoA + NAD+ enzyme FadB' converts (S)-3-hydroxybutyryl-CoA to acetoacetyl-CoA, while no conversion of (R)-3-hydroxybutyryl-CoA is detected Cupriavidus necator acetoacetyl-CoA + NADH + H+
-
 r
1.1.1.35 (S)-3-hydroxybutanoyl-CoA + NAD+
-
 Cupriavidus necator DSM 428 acetoacetyl-CoA + NADH + H+
-
 r
1.1.1.35 (S)-3-hydroxybutanoyl-CoA + NAD+ enzyme FadB' converts (S)-3-hydroxybutyryl-CoA to acetoacetyl-CoA, while no conversion of (R)-3-hydroxybutyryl-CoA is detected Cupriavidus necator DSM 428 acetoacetyl-CoA + NADH + H+
-
 r
1.1.1.35 (S)-3-hydroxybutanoyl-CoA + NADP+ relative activity of FadB' measured with NADP+ is less than 10% in comparison to the activity measured with NAD+ Cupriavidus necator acetoacetyl-CoA + NADPH + H+
-
 ?
1.1.1.35 (S)-3-hydroxybutanoyl-CoA + NADP+ relative activity of FadB' measured with NADP+ is less than 10% in comparison to the activity measured with NAD+ Cupriavidus necator DSM 428 acetoacetyl-CoA + NADPH + H+
-
 ?
1.1.1.35 additional information the N-terminal part of FadB' comprises an NAD+ binding site and is responsible for 3-hydroxyacyl-CoA dehydrogenase activity converting (S)-3-hydroxybutyryl-CoA to acetoacetyl-CoA. FadB' is strictly stereospecific to (S)-3-hydroxybutyryl-CoA and to prefers NAD+. NADP(H) is utilized at a rate of less than 10% in comparison to activity with NAD(H) Cupriavidus necator ?
-
 ?
1.1.1.35 additional information the N-terminal part of FadB' comprises an NAD+ binding site and is responsible for 3-hydroxyacyl-CoA dehydrogenase activity converting (S)-3-hydroxybutyryl-CoA to acetoacetyl-CoA. FadB' is strictly stereospecific to (S)-3-hydroxybutyryl-CoA and to prefers NAD+. NADP(H) is utilized at a rate of less than 10% in comparison to activity with NAD(H) Cupriavidus necator DSM 428 ?
-
 ?

Subunits

EC Number Subunits Comment Organism
1.1.1.35 monomer 1 * 86000, recombinant His-tagged enzme, SDS-PAGE, 1 * 85900, about, sequence calculation Cupriavidus necator
1.1.1.35 More FadB' is a polypeptide comprising a N-terminal 3HCDH_N-domain, responsible for NAD+ binding, and a C-terminal enoyl-CoA hydratase domain, an with inverted domain order Cupriavidus necator

Synonyms

EC Number Synonyms Comment Organism
1.1.1.35 (S)-3-hydroxyacyl-CoA dehydrogenase/enoyl-CoA hydratase
-
 Cupriavidus necator
1.1.1.35 (S)-stereospecific 3-hydroxyacyl-CoA dehydrogenase/enoyl-CoA hydratase
-
 Cupriavidus necator
1.1.1.35 FadB'
-
 Cupriavidus necator
1.1.1.35 H16_A0461
-
 Cupriavidus necator
1.1.1.35 More cf. EC 4.2.1.17 Cupriavidus necator

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.1.1.35 35 40
-
 Cupriavidus necator

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.1.1.35 6 7
-
 Cupriavidus necator

pH Range

EC Number pH Minimum pH Maximum Comment Organism
1.1.1.35 4 10 maximal activity at pH 6.0-7.0, 65% and 75% activity at pH 5.0 and pH 8.0, respectively, and 46% of the maximal activity at pH 10.0, and 26% of maximal activity at pH 4.0 Cupriavidus necator

Cofactor

EC Number Cofactor Comment Organism Structure
1.1.1.35 additional information the enzyme prefers NAD+. NADP(H) is utilized at a rate of less than 10% in comparison to activity with NAD(H) Cupriavidus necator
1.1.1.35 NAD+
-
 Cupriavidus necator
1.1.1.35 NADH
-
 Cupriavidus necator
1.1.1.35 NADP+
-
 Cupriavidus necator
1.1.1.35 NADPH
-
 Cupriavidus necator

pI Value

EC Number Organism Comment pI Value Maximum pI Value
1.1.1.35 Cupriavidus necator sequence calculation
-
 7.66

General Information

EC Number General Information Comment Organism
1.1.1.35 evolution two (S)-3-hydroxyacyl-CoA dehydrogenase/enoyl-CoA hydratases, H16_A0461/FadB' and H16_A1526/FadB1, are involved in the FA degradation in Ralstonia eutropha H16. FadB' and FadB1 possess an enoyl-CoA hydratase activity, catalyzing hydrogenation of the unsaturated enoyl coenzyme A (CoA), and a 3-hydroxyacyl-CoA dehydrogenase activity, i.e. oxidation of the hydroxyl group into a keto group using one NAD+ molecule Cupriavidus necator
1.1.1.35 metabolism the enzyme is involved in fatty acid degradation metabolism Cupriavidus necator
1.1.1.35 additional information FadB' is an enzyme with two catalytic domains exhibiting a single monomeric structure and possessing a molecular weight of 86 kDa. The C-terminal part of the enzyme harbors enoyl-CoA hydratase activity, EC 4.2.1.17, and is able to convert trans-crotonyl-CoA to 3-hydroxybutyryl-CoA. The N-terminal part of FadB' comprises an NAD+ binding site and is responsible for 3-hydroxyacyl-CoA dehydrogenase activity converting (S)-3-hydroxybutyryl-CoA to acetoacetyl-CoA Cupriavidus necator