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Literature summary extracted from

  • Watanabe, S.; Tozawa, Y.; Watanabe, Y.
    Ornithine cyclodeaminase/mu-crystallin homolog from the hyperthermophilic archaeon Thermococcus litoralis functions as a novel DELTA(1)-pyrroline-2-carboxylate reductase involved in putative trans-3-hydroxy-L-proline metabolism (2014), FEBS Open Bio, 4, 617-626.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.5.1.B5 His-tagged protein is expressed in Escherichia coli BL21. TlLhpI is a fusion of OCC_00362 and OCC_00367 genes by PCR. The gene from Thermococcus litoralis DSM 5473 consists of two separate open-reading frames (ORFs): OCC_00362 and OCC_00367. When a single A is inserted between A227646 and T227647 in the original genome sequence of T. litoralis DSM 5473, the separate ORFs fuse into a single ORF, and the putative amino acid sequence of is homologous to that of the Thermococcus sibiricus TSIB_0634 gene. Although there is a possibility that this OCD-like protein from Thermococcus litoralis DSM 5473 forms a heterooligomeric structure, the TlLhpI gene is synthesized as a single ORF by sequential steps of PCR using sense and antisense primers and genome DNA of Thermococcus litoralis DSM 5473 as a template Thermococcus litoralis
4.2.1.77
-
 Thermococcus litoralis

Protein Variants

EC Number Protein Variants Comment Organism
1.5.1.B5 L52E/V224D/A228K DELTA1-pyrroline-2-carboxylate reductase activity is the same as in wild-type enzyme, no alanine dehydrogenase activity Thermococcus litoralis
1.5.1.B5 L52R/V224D/A228K DELTA1-pyrroline-2-carboxylate reductase activity is the same as in wild-type enzyme, no alanine dehydrogenase activity Thermococcus litoralis
1.5.1.B5 additional information TlLhpI is a fusion of OCC_00362 and OCC_00367 genes by PCR. The gene from Thermococcus litoralis DSM 5473 consistsof two separate open-reading frames (ORFs): OCC_00362 and OCC_00367. When a single A is inserted between A227646 and T227647 in the original genome sequence of T. litoralis DSM 5473, the separate ORFs fuse into a single ORF, and the putative amino acid sequence of is homologous to that of the Thermococcus sibiricus TSIB_0634 gene. Although there is a possibility that this OCD-like protein from Thermococcus litoralis DSM 5473 forms a heterooligomeric structure, the TlLhpI gene is synthesized as a single ORF by sequential steps of PCR using sense and antisense primers and genome DNA of Thermococcus litoralis DSM 5473 as a template Thermococcus litoralis
1.5.1.B5 V224D/A228K 12fold enhancement of DELTA1-pyrroline-2-carboxylate reductase activity, 6.7fold enhancement for the oxidization activity for L-proline, no alanine dehydrogenase activity Thermococcus litoralis

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.5.1.B5 0.444
-
 L-proline pH 6.0, 50°C, mutant enzyme V224D/A228K Thermococcus litoralis
1.5.1.B5 0.608
-
 L-pipecolate pH 6.0, 50°C, wild-type enzyme Thermococcus litoralis
1.5.1.B5 0.944
-
 DELTA1-pyrroline-2-carboxylate pH 6.0, 50°C, wild-type enzyme Thermococcus litoralis
1.5.1.B5 1.12
-
 L-proline pH 6.0, 50°C, wild-type enzyme Thermococcus litoralis
1.5.1.B5 2.77
-
 DELTA1-pyrroline-2-carboxylate pH 6.0, 50°C, mutant enzyme V224D/A228K Thermococcus litoralis

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
1.5.1.B5 40000
-
 2 * 40000, SDS-PAGE Thermococcus litoralis
1.5.1.B5 72000
-
 gel filtration Thermococcus litoralis

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.5.1.B5 DELTA1-pyrroline-2-carboxylate + NADH + H+ Thermococcus litoralis the enzyme is involved in the trans-3-hydroxy-L-proline metabolism L-proline + NAD+
-
 ?
1.5.1.B5 DELTA1-pyrroline-2-carboxylate + NADH + H+ Thermococcus litoralis DSM 5473 the enzyme is involved in the trans-3-hydroxy-L-proline metabolism L-proline + NAD+
-
 ?
4.2.1.77 trans-3-hydroxy-L-proline Thermococcus litoralis the enzyme is involved in the trans-3-hydroxy-L-proline metabolism 1-pyrroline 2-carboxylate + H2O
-
 ?
4.2.1.77 trans-3-hydroxy-L-proline Thermococcus litoralis DSM 5473 the enzyme is involved in the trans-3-hydroxy-L-proline metabolism 1-pyrroline 2-carboxylate + H2O
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
1.5.1.B5 Thermococcus litoralis H3ZMH3 and H3ZMH4 TlLhpI is a fusion of OCC_00362 and OCC_00367 genes by PCR. The gene from Thermococcus litoralis DSM 5473 consists of two separate open-reading frames (ORFs): OCC_00362 and OCC_00367. When a single A is inserted between A227646 and T227647 in the original genome sequence of T. litoralis DSM 5473, the separate ORFs fuse into a single ORF, and the putative amino acid sequence of is homologous to that of the Thermococcus sibiricus TSIB_0634 gene. Although there is a possibility that this OCD-like protein from Thermococcus litoralis DSM 5473 forms a heterooligomeric structure, the TlLhpI gene is synthesized as a single ORF by sequential steps of PCR using sense and antisense primers and genome DNA of Thermococcus litoralis DSM 5473 as a template
-
1.5.1.B5 Thermococcus litoralis DSM 5473 H3ZMH3 and H3ZMH4 TlLhpI is a fusion of OCC_00362 and OCC_00367 genes by PCR. The gene from Thermococcus litoralis DSM 5473 consists of two separate open-reading frames (ORFs): OCC_00362 and OCC_00367. When a single A is inserted between A227646 and T227647 in the original genome sequence of T. litoralis DSM 5473, the separate ORFs fuse into a single ORF, and the putative amino acid sequence of is homologous to that of the Thermococcus sibiricus TSIB_0634 gene. Although there is a possibility that this OCD-like protein from Thermococcus litoralis DSM 5473 forms a heterooligomeric structure, the TlLhpI gene is synthesized as a single ORF by sequential steps of PCR using sense and antisense primers and genome DNA of Thermococcus litoralis DSM 5473 as a template
-
4.2.1.77 Thermococcus litoralis H3ZMH8
-
-
4.2.1.77 Thermococcus litoralis DSM 5473 H3ZMH8
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.5.1.B5
-
 Thermococcus litoralis
4.2.1.77
-
 Thermococcus litoralis

Reaction

EC Number Reaction Comment Organism Reaction ID
1.5.1.B5 L-pipecolate + NAD+ = DELTA1-piperideine-2-carboxylate + NADH + H+ (1) Thermococcus litoralis
a
1.5.1.B5 L-proline + NAD+ = DELTA1-pyrroline-2-carboxylate + NADH + H+ (2) Thermococcus litoralis
a

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.5.1.B5 cis-4-hydroxy-L-proline + NAD+ activity is 85% compared to the activity with L-proline Thermococcus litoralis ?
-
 ?
1.5.1.B5 cis-4-hydroxy-L-proline + NAD+ activity is 85% compared to the activity with L-proline Thermococcus litoralis DSM 5473 ?
-
 ?
1.5.1.B5 DELTA1-pyrroline-2-carboxylate + NADH + H+ the enzyme is involved in the trans-3-hydroxy-L-proline metabolism Thermococcus litoralis L-proline + NAD+
-
 ?
1.5.1.B5 DELTA1-pyrroline-2-carboxylate + NADH + H+ preference of the reaction equilibrium in the direction toward NADH-dependent reduction Thermococcus litoralis L-proline + NAD+
-
 r
1.5.1.B5 DELTA1-pyrroline-2-carboxylate + NADH + H+ the enzyme is involved in the trans-3-hydroxy-L-proline metabolism Thermococcus litoralis DSM 5473 L-proline + NAD+
-
 ?
1.5.1.B5 DELTA1-pyrroline-2-carboxylate + NADH + H+ preference of the reaction equilibrium in the direction toward NADH-dependent reduction Thermococcus litoralis DSM 5473 L-proline + NAD+
-
 r
1.5.1.B5 L-pipecolate + NAD+ activity is 49% compared to the activity with L-proline Thermococcus litoralis DELTA1-piperideine-2-carboxylate + NADH + H+
-
 ?
1.5.1.B5 L-pipecolate + NAD+ activity is 49% compared to the activity with L-proline Thermococcus litoralis DSM 5473 DELTA1-piperideine-2-carboxylate + NADH + H+
-
 ?
1.5.1.B5 L-proline + NAD+ preference of the reaction equilibrium in the direction toward NADH-dependent reduction Thermococcus litoralis DELTA1-pyrroline-2-carboxylate + NADH + H+
-
 r
1.5.1.B5 additional information the enzyme also shows alanine dehydrogenase activity (EC 1.4.1.1) and N-methyl-L-alanine dehydrogenase activity (EC 1.4.1.17) Thermococcus litoralis ?
-
 ?
1.5.1.B5 additional information the enzyme also shows alanine dehydrogenase activity (EC 1.4.1.1) and N-methyl-L-alanine dehydrogenase activity (EC 1.4.1.17) Thermococcus litoralis DSM 5473 ?
-
 ?
1.5.1.B5 trans-4-hydroxy-L-proline + NAD+ activity is 70% compared to the activity with L-proline Thermococcus litoralis ?
-
 ?
4.2.1.77 trans-3-hydroxy-L-proline
-
 Thermococcus litoralis 1-pyrroline 2-carboxylate + H2O
-
 ?
4.2.1.77 trans-3-hydroxy-L-proline the enzyme is involved in the trans-3-hydroxy-L-proline metabolism Thermococcus litoralis 1-pyrroline 2-carboxylate + H2O
-
 ?
4.2.1.77 trans-3-hydroxy-L-proline
-
 Thermococcus litoralis DSM 5473 1-pyrroline 2-carboxylate + H2O
-
 ?
4.2.1.77 trans-3-hydroxy-L-proline the enzyme is involved in the trans-3-hydroxy-L-proline metabolism Thermococcus litoralis DSM 5473 1-pyrroline 2-carboxylate + H2O
-
 ?

Subunits

EC Number Subunits Comment Organism
1.5.1.B5 dimer 2 * 40000, SDS-PAGE Thermococcus litoralis

Synonyms

EC Number Synonyms Comment Organism
1.5.1.B5 DELTA1-pyrroline-2-carboxylate reductase misleading Thermococcus litoralis
1.5.1.B5 Pyr2C reductase
-
 Thermococcus litoralis
1.5.1.B5 TlLhpI
-
 Thermococcus litoralis
4.2.1.77 T3LHyp dehydratase
-
 Thermococcus litoralis
4.2.1.77 TlLhpH
-
 Thermococcus litoralis

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.5.1.B5 50
-
 assay at Thermococcus litoralis
4.2.1.77 80 100
-
 Thermococcus litoralis

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.5.1.B5 0.2
-
 L-pipecolate pH 6.0, 50°C, wild-type enzyme Thermococcus litoralis
1.5.1.B5 0.23
-
 L-proline pH 6.0, 50°C, wild-type enzyme Thermococcus litoralis
1.5.1.B5 0.61
-
 L-proline pH 6.0, 50°C, mutant enzyme V224D/A228K Thermococcus litoralis
1.5.1.B5 16.6
-
 DELTA1-pyrroline-2-carboxylate pH 6.0, 50°C, wild-type enzyme Thermococcus litoralis
1.5.1.B5 558
-
 DELTA1-pyrroline-2-carboxylate pH 6.0, 50°C, mutant enzyme V224D/A228K Thermococcus litoralis

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.5.1.B5 4 6 substrates: DELTA1-pyrroline-2-carboxylate + NADH + H+ Thermococcus litoralis
1.5.1.B5 11
-
 substrates: L-proline + NAD+ Thermococcus litoralis

pH Range

EC Number pH Minimum pH Maximum Comment Organism
1.5.1.B5 5 11 pH 5.0: about 40% of maximal activity, pH 11.0: about 80% of maximal activity Thermococcus litoralis
1.5.1.B5 10 12 pH 10.0: about 75% of maximal activity, pH 12.0: about 85% of maximal activity Thermococcus litoralis

Cofactor

EC Number Cofactor Comment Organism Structure
1.5.1.B5 NAD+ the kcat/Km value for proline in the presence of NADP+ is about 50fold lower than that in the presence of NAD+ Thermococcus litoralis
1.5.1.B5 NADH no activity with NADPH Thermococcus litoralis

General Information

EC Number General Information Comment Organism
1.5.1.B5 metabolism the enzyme is involved in the trans-3-hydroxy-L-proline metabolism Thermococcus litoralis
4.2.1.77 metabolism the enzyme is involved in the trans-3-hydroxy-L-proline metabolism Thermococcus litoralis

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
1.5.1.B5 0.2
-
 L-proline pH 6.0, 50°C, wild-type enzyme Thermococcus litoralis
1.5.1.B5 0.33
-
 L-pipecolate pH 6.0, 50°C, wild-type enzyme Thermococcus litoralis
1.5.1.B5 1.4
-
 L-proline pH 6.0, 50°C, mutant enzyme V224D/A228K Thermococcus litoralis
1.5.1.B5 17.6
-
 DELTA1-pyrroline-2-carboxylate pH 6.0, 50°C, wild-type enzyme Thermococcus litoralis
1.5.1.B5 201
-
 DELTA1-pyrroline-2-carboxylate pH 6.0, 50°C, mutant enzyme V224D/A228K Thermococcus litoralis