Cloned (Comment) | Organism |
---|---|
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 |
Protein Variants | Comment | Organism |
---|---|---|
L52E/V224D/A228K | DELTA1-pyrroline-2-carboxylate reductase activity is the same as in wild-type enzyme, no alanine dehydrogenase activity | Thermococcus litoralis |
L52R/V224D/A228K | DELTA1-pyrroline-2-carboxylate reductase activity is the same as in wild-type enzyme, no alanine dehydrogenase activity | Thermococcus litoralis |
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 |
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] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.444 | - |
L-proline | pH 6.0, 50°C, mutant enzyme V224D/A228K | Thermococcus litoralis | |
0.608 | - |
L-pipecolate | pH 6.0, 50°C, wild-type enzyme | Thermococcus litoralis | |
0.944 | - |
DELTA1-pyrroline-2-carboxylate | pH 6.0, 50°C, wild-type enzyme | Thermococcus litoralis | |
1.12 | - |
L-proline | pH 6.0, 50°C, wild-type enzyme | Thermococcus litoralis | |
2.77 | - |
DELTA1-pyrroline-2-carboxylate | pH 6.0, 50°C, mutant enzyme V224D/A228K | Thermococcus litoralis |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
40000 | - |
2 * 40000, SDS-PAGE | Thermococcus litoralis |
72000 | - |
gel filtration | Thermococcus litoralis |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
DELTA1-pyrroline-2-carboxylate + NADH + H+ | Thermococcus litoralis | the enzyme is involved in the trans-3-hydroxy-L-proline metabolism | L-proline + NAD+ | - |
? | |
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+ | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
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 | - |
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 | - |
Purification (Comment) | Organism |
---|---|
- |
Thermococcus litoralis |
Reaction | Comment | Organism | Reaction ID |
---|---|---|---|
L-pipecolate + NAD+ = DELTA1-piperideine-2-carboxylate + NADH + H+ | (1) | Thermococcus litoralis | |
L-proline + NAD+ = DELTA1-pyrroline-2-carboxylate + NADH + H+ | (2) | Thermococcus litoralis |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
cis-4-hydroxy-L-proline + NAD+ | activity is 85% compared to the activity with L-proline | Thermococcus litoralis | ? | - |
? | |
cis-4-hydroxy-L-proline + NAD+ | activity is 85% compared to the activity with L-proline | Thermococcus litoralis DSM 5473 | ? | - |
? | |
DELTA1-pyrroline-2-carboxylate + NADH + H+ | the enzyme is involved in the trans-3-hydroxy-L-proline metabolism | Thermococcus litoralis | L-proline + NAD+ | - |
? | |
DELTA1-pyrroline-2-carboxylate + NADH + H+ | preference of the reaction equilibrium in the direction toward NADH-dependent reduction | Thermococcus litoralis | L-proline + NAD+ | - |
r | |
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+ | - |
? | |
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 | |
L-pipecolate + NAD+ | activity is 49% compared to the activity with L-proline | Thermococcus litoralis | DELTA1-piperideine-2-carboxylate + NADH + H+ | - |
? | |
L-pipecolate + NAD+ | activity is 49% compared to the activity with L-proline | Thermococcus litoralis DSM 5473 | DELTA1-piperideine-2-carboxylate + NADH + H+ | - |
? | |
L-proline + NAD+ | preference of the reaction equilibrium in the direction toward NADH-dependent reduction | Thermococcus litoralis | DELTA1-pyrroline-2-carboxylate + NADH + H+ | - |
r | |
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 | ? | - |
? | |
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 | ? | - |
? | |
trans-4-hydroxy-L-proline + NAD+ | activity is 70% compared to the activity with L-proline | Thermococcus litoralis | ? | - |
? |
Subunits | Comment | Organism |
---|---|---|
dimer | 2 * 40000, SDS-PAGE | Thermococcus litoralis |
Synonyms | Comment | Organism |
---|---|---|
DELTA1-pyrroline-2-carboxylate reductase | misleading | Thermococcus litoralis |
Pyr2C reductase | - |
Thermococcus litoralis |
TlLhpI | - |
Thermococcus litoralis |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
50 | - |
assay at | Thermococcus litoralis |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.2 | - |
L-pipecolate | pH 6.0, 50°C, wild-type enzyme | Thermococcus litoralis | |
0.23 | - |
L-proline | pH 6.0, 50°C, wild-type enzyme | Thermococcus litoralis | |
0.61 | - |
L-proline | pH 6.0, 50°C, mutant enzyme V224D/A228K | Thermococcus litoralis | |
16.6 | - |
DELTA1-pyrroline-2-carboxylate | pH 6.0, 50°C, wild-type enzyme | Thermococcus litoralis | |
558 | - |
DELTA1-pyrroline-2-carboxylate | pH 6.0, 50°C, mutant enzyme V224D/A228K | Thermococcus litoralis |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
4 | 6 | substrates: DELTA1-pyrroline-2-carboxylate + NADH + H+ | Thermococcus litoralis |
11 | - |
substrates: L-proline + NAD+ | Thermococcus litoralis |
pH Minimum | pH Maximum | Comment | Organism |
---|---|---|---|
5 | 11 | pH 5.0: about 40% of maximal activity, pH 11.0: about 80% of maximal activity | Thermococcus litoralis |
10 | 12 | pH 10.0: about 75% of maximal activity, pH 12.0: about 85% of maximal activity | Thermococcus litoralis |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
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 | |
NADH | no activity with NADPH | Thermococcus litoralis |
General Information | Comment | Organism |
---|---|---|
metabolism | the enzyme is involved in the trans-3-hydroxy-L-proline metabolism | Thermococcus litoralis |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.2 | - |
L-proline | pH 6.0, 50°C, wild-type enzyme | Thermococcus litoralis | |
0.33 | - |
L-pipecolate | pH 6.0, 50°C, wild-type enzyme | Thermococcus litoralis | |
1.4 | - |
L-proline | pH 6.0, 50°C, mutant enzyme V224D/A228K | Thermococcus litoralis | |
17.6 | - |
DELTA1-pyrroline-2-carboxylate | pH 6.0, 50°C, wild-type enzyme | Thermococcus litoralis | |
201 | - |
DELTA1-pyrroline-2-carboxylate | pH 6.0, 50°C, mutant enzyme V224D/A228K | Thermococcus litoralis |