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Literature summary for 4.2.1.148 extracted from

  • Borjian, F.; Johnsen, U.; Schoenheit, P.; Berg, I.A.
    Succinyl-CoA mesaconate CoA-transferase and mesaconyl-CoA hydratase, enzymes of the methylaspartate cycle in Haloarcula hispanica (2017), Front. Microbiol., 8, 1683 .
    View publication on PubMedView publication on EuropePMC
Search for more literature for 4.2.1.148

Cloned(Commentary)

Cloned (Comment) Organism
gene Hah_1340, phylogenetic analysis, recombinant expression of soluble His-tagged enzyme in Haloferax volcanii strain H1209 Haloarcula hispanica
Haloferax volcanii strain H1209 is used for the heterologous expression of the Haloarcula hispanica proteins Haloarcula hispanica

Inhibitors

Inhibitors Comment Organism Structure
CaCl2 18% inhibition at 5 mM; 5 mM, 18% inhibition Haloarcula hispanica
CoCl2 40% inhibition at 5 mM; 5 mM, 34% inhibition Haloarcula hispanica
MgCl2 11% inhibition at 5 mM; 5 mM, 11% inhibition Haloarcula hispanica
MnCl2 22% inhibition at 5 mM; 5 mM, 22% inhibition Haloarcula hispanica
additional information no inhibition by EDTA; no inhibition by EDTA at 2 mM Haloarcula hispanica
NiCl2 34% inhibition at 5 mM; 5 mM, 40% inhibition Haloarcula hispanica

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.18
-
 3-methylfumaryl-CoA 37°C, pH 7.8 Haloarcula hispanica
0.32
-
 (S)-malyl-CoA 37°C, pH 7.8 Haloarcula hispanica
0.35
-
 (2R,3S)-2-methylmalyl-CoA 37°C, pH 7.8 Haloarcula hispanica
1
-
 2-methylfumaryl-CoA 37°C, pH 7.8 Haloarcula hispanica
2.6
-
 (S)-citramalyl-CoA 37°C, pH 7.8 Haloarcula hispanica

Metals/Ions

Metals/Ions Comment Organism Structure
KCl the enzyme is dependent on the presence of KCl and shows highest activity at 0.5 M KCl. With 82% of activity at 4 M KCl, whereas the activity dropps severely at 0.1 M KCl Haloarcula hispanica
additional information no requirement for divalent cations Haloarcula hispanica

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
86000
-
 gel filtration Haloarcula hispanica
86000
-
 recombinant His-tagged enzyme, gel filtration Haloarcula hispanica

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2-methylfumaryl-CoA + H2O Haloarcula hispanica i.e. mesaconyl-CoA. The enzyme catalyzes a reaction in the methylaspartate cycle (2R,3S)-2-methylmalyl-CoA
-
 ?
2-methylfumaryl-CoA + H2O Haloarcula hispanica ATCC 33960 / DSM 4426 / JCM 8911 / NBRC 102182 / NCIMB 2187 / VKM B-1755 i.e. mesaconyl-CoA. The enzyme catalyzes a reaction in the methylaspartate cycle (2R,3S)-2-methylmalyl-CoA
-
 ?
2-methylfumaryl-CoA + H2O Haloarcula hispanica DSM 4426 i.e. mesaconyl-CoA. The enzyme catalyzes a reaction in the methylaspartate cycle (2R,3S)-2-methylmalyl-CoA
-
 ?
mesaconyl-C1-CoA Haloarcula hispanica the physiological substrate is mesaconyl-C1-CoA (2R,3S)-2-methyl-3-hydroxysuccinyl-C1-CoA + H2O
-
 r
mesaconyl-C1-CoA Haloarcula hispanica ATCC 33960 / DSM 4426 / JCM 8911 / NBRC 102182 / NCIMB 2187 / VKM B-1755 the physiological substrate is mesaconyl-C1-CoA (2R,3S)-2-methyl-3-hydroxysuccinyl-C1-CoA + H2O
-
 r
mesaconyl-C1-CoA Haloarcula hispanica DSM 4426 the physiological substrate is mesaconyl-C1-CoA (2R,3S)-2-methyl-3-hydroxysuccinyl-C1-CoA + H2O
-
 r

Organism

Organism UniProt Comment Textmining
Haloarcula hispanica G0HQ35
-
-
Haloarcula hispanica ATCC 33960 / DSM 4426 / JCM 8911 / NBRC 102182 / NCIMB 2187 / VKM B-1755 G0HQ35
-
-
Haloarcula hispanica DSM 4426 G0HQ35
-
-

Purification (Commentary)

Purification (Comment) Organism
-
 Haloarcula hispanica
recombinant His-tagged enzyme 427fold from Haloferax volcanii strain H1209 by nickel affinity chromatography and gel filtration Haloarcula hispanica

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
157
-
 37°C, pH 7.8 Haloarcula hispanica
157
-
 purified recombinant His-tagged enzyme, substrate mesaconyl-C1-CoA, pH 7.8, 37°C Haloarcula hispanica

Storage Stability

Storage Stability Organism
-20°C, purified recombinant enzyme, 50 mM Tris/HCl, pH 8.0, 2 M KCl, 3 weeks, complete loss of activity Haloarcula hispanica
4°C, purified recombinant enzyme, 50 mM Tris/HCl, pH 8.0, 2 M KCl, 3 weeks, loss of 70-80% activity Haloarcula hispanica

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
(2R,3S)-2-methyl-3-hydroxysuccinyl-C4-CoA + H2O i.e. (S)-citramalyl-CoA Haloarcula hispanica 3-methylfumaryl-CoA + H2O
-
 r
(2R,3S)-2-methylmalyl-CoA the equilibrium of the reaction is on the side of (2R,3S)-2-methylmalyl-CoA Haloarcula hispanica 2-methylfumaryl-CoA + H2O
-
 r
(2R,3S)-2-methylmalyl-CoA the equilibrium of the reaction is on the side of (2R,3S)-2-methylmalyl-CoA Haloarcula hispanica ATCC 33960 / DSM 4426 / JCM 8911 / NBRC 102182 / NCIMB 2187 / VKM B-1755 2-methylfumaryl-CoA + H2O
-
 r
(2R,3S)-2-methylmalyl-CoA the equilibrium of the reaction is on the side of (2R,3S)-2-methylmalyl-CoA Haloarcula hispanica DSM 4426 2-methylfumaryl-CoA + H2O
-
 r
(S)-citramalyl-CoA
-
 Haloarcula hispanica 3-methylfumaryl-CoA + H2O
-
 r
(S)-malyl-CoA
-
 Haloarcula hispanica fumaryl-CoA + H2O
-
 r
2-methylfumaryl-CoA + H2O i.e. mesaconyl-CoA. The enzyme catalyzes a reaction in the methylaspartate cycle Haloarcula hispanica (2R,3S)-2-methylmalyl-CoA
-
 ?
2-methylfumaryl-CoA + H2O i.e. mesaconyl-CoA, i.e. mesaconyl-C1-CoA. The equilibrium of the reaction is on the side of (2R,3S)-2-methylmalyl-CoA Haloarcula hispanica (2R,3S)-2-methylmalyl-CoA
-
 r
2-methylfumaryl-CoA + H2O i.e. mesaconyl-CoA. The enzyme catalyzes a reaction in the methylaspartate cycle Haloarcula hispanica ATCC 33960 / DSM 4426 / JCM 8911 / NBRC 102182 / NCIMB 2187 / VKM B-1755 (2R,3S)-2-methylmalyl-CoA
-
 ?
2-methylfumaryl-CoA + H2O i.e. mesaconyl-CoA, i.e. mesaconyl-C1-CoA. The equilibrium of the reaction is on the side of (2R,3S)-2-methylmalyl-CoA Haloarcula hispanica ATCC 33960 / DSM 4426 / JCM 8911 / NBRC 102182 / NCIMB 2187 / VKM B-1755 (2R,3S)-2-methylmalyl-CoA
-
 r
2-methylfumaryl-CoA + H2O i.e. mesaconyl-CoA. The enzyme catalyzes a reaction in the methylaspartate cycle Haloarcula hispanica DSM 4426 (2R,3S)-2-methylmalyl-CoA
-
 ?
2-methylfumaryl-CoA + H2O i.e. mesaconyl-CoA, i.e. mesaconyl-C1-CoA. The equilibrium of the reaction is on the side of (2R,3S)-2-methylmalyl-CoA Haloarcula hispanica DSM 4426 (2R,3S)-2-methylmalyl-CoA
-
 r
3-methylfumaryl-CoA + H2O i.e. mesaconyl-C4-CoA Haloarcula hispanica (S)-citramalyl-CoA
-
 r
fumaryl-CoA + H2O
-
 Haloarcula hispanica (S)-malyl-CoA
-
 r
mesaconyl-C1-CoA the physiological substrate is mesaconyl-C1-CoA Haloarcula hispanica (2R,3S)-2-methyl-3-hydroxysuccinyl-C1-CoA + H2O
-
 r
mesaconyl-C1-CoA hydration of mesaconyl-C1-CoA to (2R,3S)-2-methyl-3-hydroxysuccinyl-C1-CoA (erythro-beta-methylmalyl-CoA), EC 4.2.1.148. The activity for hydration of mesaconyl-C1-CoA is an order of magnitude higher for the reverse reaction of beta-methylmalyl-CoA dehydration Haloarcula hispanica (2R,3S)-2-methyl-3-hydroxysuccinyl-C1-CoA + H2O
-
 r
mesaconyl-C1-CoA the physiological substrate is mesaconyl-C1-CoA Haloarcula hispanica ATCC 33960 / DSM 4426 / JCM 8911 / NBRC 102182 / NCIMB 2187 / VKM B-1755 (2R,3S)-2-methyl-3-hydroxysuccinyl-C1-CoA + H2O
-
 r
mesaconyl-C1-CoA the physiological substrate is mesaconyl-C1-CoA Haloarcula hispanica DSM 4426 (2R,3S)-2-methyl-3-hydroxysuccinyl-C1-CoA + H2O
-
 r
mesaconyl-C4-CoA + H2O EC 4.2.1.153 Haloarcula hispanica (2R,3S)-2-methyl-3-hydroxysuccinyl-C4-CoA + H2O
-
 r
additional information recombinant mesaconyl-CoA hydratase has a 3.5fold bias for the physiological substrate, mesaconyl-C1-CoA, compared to mesaconyl-C4-CoA. The enzyme is also active with mesaconyl-C4-CoA, (S)-citramalyl-CoA and (S)-malyl-CoA, and virtually has no activity with other tested enoyl-CoA/3-hydroxyacyl-CoA compounds. The enzyme activity is an order of magnitude higher for mesaconyl-C1-CoA/beta-methylmalyl-CoA compared to other substrates Haloarcula hispanica ?
-
 ?
additional information recombinant mesaconyl-CoA hydratase has a 3.5fold bias for the physiological substrate, mesaconyl-C1-CoA, compared to mesaconyl-C4-CoA. The enzyme is also active with mesaconyl-C4-CoA, (S)-citramalyl-CoA and (S)-malyl-CoA, and virtually has no activity with other tested enoyl-CoA/3-hydroxyacyl-CoA compounds. The enzyme activity is an order of magnitude higher for mesaconyl-C1-CoA/beta-methylmalyl-CoA compared to other substrates Haloarcula hispanica ATCC 33960 / DSM 4426 / JCM 8911 / NBRC 102182 / NCIMB 2187 / VKM B-1755 ?
-
 ?
additional information recombinant mesaconyl-CoA hydratase has a 3.5fold bias for the physiological substrate, mesaconyl-C1-CoA, compared to mesaconyl-C4-CoA. The enzyme is also active with mesaconyl-C4-CoA, (S)-citramalyl-CoA and (S)-malyl-CoA, and virtually has no activity with other tested enoyl-CoA/3-hydroxyacyl-CoA compounds. The enzyme activity is an order of magnitude higher for mesaconyl-C1-CoA/beta-methylmalyl-CoA compared to other substrates Haloarcula hispanica DSM 4426 ?
-
 ?

Subunits

Subunits Comment Organism
dimer 2 * 43000, recombinant His-tagged enzyme, SDS-PAGE, 2 * 41000, about, sequence calculation Haloarcula hispanica
homodimer 2 * 43000, SDS-PAGE Haloarcula hispanica
More haloarchaeal Mch consists of two (R)-specific enoyl-CoA hydratase domains Haloarcula hispanica

Synonyms

Synonyms Comment Organism
Hah_1340
-
 Haloarcula hispanica
Mch
-
 Haloarcula hispanica
mesaconyl-CoA hydratase
-
 Haloarcula hispanica
More cf. EC 4.2.1.153 Haloarcula hispanica

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
 assay at Haloarcula hispanica

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.8
-
 assay at Haloarcula hispanica

pI Value

Organism Comment pI Value Maximum pI Value
Haloarcula hispanica calculated from sequence
-
 4.7

General Information

General Information Comment Organism
evolution the mch gene is mainly transmitted vertically among haloarchaea Haloarcula hispanica
evolution the methylaspartate cycle is discovered in Haloarcula spp. and operates in about 40% of sequenced haloarchaea. Mesaconate CoA-transferase and mesaconyl-CoA hydratase are the characteristic enzymes of this cycle. Phylogenetic analysis Haloarcula hispanica
metabolism mesaconate (methylfumarate) that needs to be activated to mesaconyl-C1-CoA and hydrated to beta-methylmalyl-CoA in the methylaspartate cycle as part of the anaplerotic pathway for the conversion of acetyl-CoA into cellular building blocks. The cleavage of beta-methylmalyl-CoA results in the formation of propionyl-CoA and glyoxylate. The carboxylation of propionyl-CoA and the condensation of glyoxylate with another acetyl-CoA molecule give rise to two C4-dicarboxylic acids, thus regenerating the initial acetyl-CoA acceptor and forming malate, its final product. The methylaspartate cycle, overview Haloarcula hispanica
metabolism the enzyme catalyzes a reaction in the methylaspartate cycle Haloarcula hispanica

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.2
-
 (S)-citramalyl-CoA 37°C, pH 7.8 Haloarcula hispanica
2
-
 3-methylfumaryl-CoA 37°C, pH 7.8 Haloarcula hispanica
3.8
-
 (S)-malyl-CoA 37°C, pH 7.8 Haloarcula hispanica
6.9
-
 2-methylfumaryl-CoA 37°C, pH 7.8 Haloarcula hispanica
254
-
 (2R,3S)-2-methylmalyl-CoA 37°C, pH 7.8 Haloarcula hispanica