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Literature summary extracted from
- An enzymatic method to estimate the content of L-hydroxyproline (2015), J. Biotechnol., 199, 9-16 .
Application
| EC Number | Application | Comment | Organism |
|---|---|---|---|
| 1.4.99.B4 | analysis | estimation of the content of L-hydroxyprolines using coupling systems with metabolic enzymes of the trans-4-hydroxy-L-proline pathway (hydroxyproline 2-epimerase (HypE) and cis-4-hydroxy-D-proline dehydrogenase (HypDH)) and the trans-3-hydroxy-L-proline pathway (trans-3-hydroxy-L-proline dehydratase (T3LHypD) and DELTA1-pyrroline-2-carboxylate reductase (Pyr2CR)) from microorganisms. A functional expression system of recombinant HypDH with a heterooligomeric structure is constructed in Escherichia coli cells. Enzymological characterization reveals that the beta-subunit acts as a catalytic subunit, and also that assembly with other subunit(s) improves the kinetics for cis-4-hydroxy-D-proline and thermostability. By using a spectrophotometric assay with different wavelengths, the contents of trans-4-hydroxy-L-proline and trans-3-hydroxy-L-proline are successfully estimated within the ranges of 0.004-1 mM and 0.05-1 mM, respectively, and are consistent with the contents determined by HPLC. This enzymatic method is used to measure the content of trans-4-hydroxy-L-proline in the acid-hydrolysate of collagen, and blood plasma | Azospirillum brasilense |
| 4.2.1.77 | analysis | estimation of the content of L-hydroxyprolines using coupling systems with metabolic enzymes of the trans-4-hydroxy-L-proline pathway (hydroxyproline 2-epimerase (HypE) and cis-4-hydroxy-D-proline dehydrogenase (HypDH)) and the trans-3-hydroxy-L-proline pathway (trans-3-hydroxy-L-proline dehydratase (T3LHypD) and DELTA1-pyrroline-2-carboxylate reductase (Pyr2CR)) from microorganisms. A functional expression system of recombinant HypDH with a heterooligomeric structure is constructed in Escherichia coli cells. Enzymological characterization reveals that the beta-subunit acts as a catalytic subunit, and also that assembly with other subunit(s) improves the kinetics for cis-4-hydroxy-D-proline and thermostability. By using a spectrophotometric assay with different wavelengths, the contents of trans-4-hydroxy-L-proline and trans-3-hydroxy-L-proline are successfully estimated within the ranges of 0.004-1 mM and 0.05-1 mM, respectively, and are consistent with the contents determined by HPLC. This enzymatic method is used to measure the content of trans-4-hydroxy-L-proline in the acid-hydrolysate of collagen, and blood plasma | Colwellia psychrerythraea |
| 4.2.1.77 | analysis | estimation of the content of L-hydroxyprolines using coupling systems with metabolic enzymes of the trans-4-hydroxy-L-proline pathway (hydroxyproline 2-epimerase (HypE) and cis-4-hydroxy-D-proline dehydrogenase (HypDH)) and the trans-3-hydroxy-L-proline pathway (trans-3-hydroxy-L-proline dehydratase (T3LHypD) and DELTA1-pyrroline-2-carboxylate reductase (Pyr2CR)) from microorganisms. A functional expression system of recombinant HypDH with a heterooligomeric structure is constructed in Escherichia coli cells. Enzymological characterization reveals that the beta-subunit acts as a catalytic subunit, and also that assembly with other subunit(s) improves the kinetics for cis-4-hydroxy-D-proline and thermostability. By using a spectrophotometric assay with different wavelengths, the contents of trans-4-hydroxy-L-proline and trans-3-hydroxy-L-proline are successfully estimated within the ranges of 0.004-1 mM and 0.05-1 mM, respectively, and are consistent with the contents determined by HPLC. This enzymatic method is used to measure the content of trans-4-hydroxy-L-proline in the acid-hydrolysate of collagen, and blood plasma | Azospirillum brasilense |
| 4.2.1.77 | analysis | estimation of the content of L-hydroxyprolines using coupling systems with metabolic enzymes of the trans-4-hydroxy-L-proline pathway (hydroxyproline 2-epimerase (HypE) and cis-4-hydroxy-D-proline dehydrogenase (HypDH)) and the trans-3-hydroxy-L-proline pathway (trans-3-hydroxy-L-proline dehydratase (T3LHypD) and DELTA1-pyrroline-2-carboxylate reductase (Pyr2CR)) from microorganisms. A functional expression system of recombinant HypDH with a heterooligomeric structure is constructed in Escherichia coli cells. Enzymological characterization reveals that the beta-subunit acts as a catalytic subunit, and also that assembly with other subunit(s) improves the kinetics for cis-4-hydroxy-D-proline and thermostability. By using a spectrophotometric assay with different wavelengths, the contents of trans-4-hydroxy-L-proline and trans-3-hydroxy-L-proline are successfully estimated within the ranges of 0.004-1 mM and 0.05-1 mM, respectively, and are consistent with the contents determined by HPLC. This enzymatic method is used to measure the content of trans-4-hydroxy-L-proline in the acid-hydrolysate of collagen, and blood plasma | Homo sapiens |
| 5.1.1.8 | analysis | estimation of the content of L-hydroxyprolines using coupling systems with metabolic enzymes of the trans-4-hydroxy-L-proline pathway (hydroxyproline 2-epimerase (HypE) and cis-4-hydroxy-D-proline dehydrogenase (HypDH)) and the trans-3-hydroxy-L-proline pathway (trans-3-hydroxy-L-proline dehydratase (T3LHypD) and DELTA1-pyrroline-2-carboxylate reductase (Pyr2CR)) from microorganisms. A functional expression system of recombinant HypDH with a heterooligomeric structure is constructed in Escherichia coli cells. Enzymological characterization reveals that the beta-subunit acts as a catalytic subunit, and also that assembly with other subunit(s) improves the kinetics for cis-4-hydroxy-D-proline and thermostability. By using a spectrophotometric assay with different wavelengths, the contents of trans-4-hydroxy-L-proline and trans-3-hydroxy-L-proline are successfully estimated within the ranges of 0.004-1 mM and 0.05-1 mM, respectively, and are consistent with the contents determined by HPLC. This enzymatic method is used to measure the content of trans-4-hydroxy-L-proline in the acid-hydrolysate of collagen, and blood plasma | Azospirillum brasilense |
Cloned(Commentary)
| EC Number | Cloned (Comment) | Organism |
|---|---|---|
| 1.4.99.B4 | cloning of the AbLhpB, AbLhpE and AbLhpF genes into different plasmid vectors, construction of a functional expression system of recombinant HypDH with a heterooligomeric structure in Escherichia coli cells | Azospirillum brasilense |
KM Value [mM]
| EC Number | KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|---|
| 1.4.99.B4 | 0.00814 | - |
trans-4-hydroxy-D-proline | pH 8.0, 30°C, AbLhpBEF protein | Azospirillum brasilense |  |
| 1.4.99.B4 | 0.0224 | - |
D-proline | pH 8.0, 30°C, AbLhpBEF protein | Azospirillum brasilense | |
| 1.4.99.B4 | 0.0628 | - |
cis-4-hydroxy-D-proline | pH 8.0, 30°C, AbLhpBEF protein | Azospirillum brasilense | |
| 4.2.1.77 | 2.79 | - |
trans-3-hydroxy-L-proline | pH 8.0, 30°C | Colwellia psychrerythraea | |
| 4.2.1.77 | 9.34 | - |
trans-3-hydroxy-L-proline | pH 8.0, 30°C | Homo sapiens | |
| 4.2.1.77 | 14.3 | - |
trans-3-hydroxy-L-proline | pH 8.0, 30°C | Azospirillum brasilense | |
Natural Substrates/ Products (Substrates)
| EC Number | Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|---|
| 1.4.99.B4 | cis-4-hydroxy-D-proline + acceptor | Azospirillum brasilense | the enzyme is involved in the metabolic pathway of trans-4-hydroxy-L-proline | (4R)-DELTA1-pyrroline-4-hydroxy-2-carboxylate + reduced acceptor | - |
? | |
| 5.1.1.8 | trans-4-hydroxy-L-proline | Azospirillum brasilense | no activity with L-proline | cis-4-hydroxy-D-proline | - |
? | |
Organism
| EC Number | Organism | UniProt | Comment | Textmining |
|---|---|---|---|---|
| 1.4.99.B4 | Azospirillum brasilense | S6C8W7 AND S6CFJ1 AND S6CLS6 | S6C8W7: beta-subunit (catalytic subunit, AbLhpB), S6CFJ1: alpha-subunit (AbLhpE), S6CLS6: gamma-subunit (AbLhpF) | - |
| 4.2.1.77 | Azospirillum brasilense | V5YXI5 | - |
- |
| 4.2.1.77 | Colwellia psychrerythraea | Q485S0 | - |
- |
| 4.2.1.77 | Homo sapiens | Q96EM0 | - |
- |
| 5.1.1.8 | Azospirillum brasilense | S6CGK1 | - |
- |
Specific Activity [micromol/min/mg]
| EC Number | Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
|---|---|---|---|---|
| 1.4.99.B4 | 1.02 | - |
substrate: D-proline, pH 8.0, 30°C, AbLhpBEF protein | Azospirillum brasilense |
| 1.4.99.B4 | 2.98 | - |
substrate: cis-4-hydroxy-D-proline, pH 8.0, 30°C, AbLhpBEF protein | Azospirillum brasilense |
| 4.2.1.77 | 7.37 | - |
substrate: trans-3-hydroxy-L-proline, 30°C, pH 8.0 | Homo sapiens |
| 4.2.1.77 | 25.4 | - |
substrate: trans-3-hydroxy-L-proline, 30°C, pH 8.0 | Colwellia psychrerythraea |
| 4.2.1.77 | 26.1 | - |
substrate: trans-3-hydroxy-L-proline, 30°C, pH 8.0 | Azospirillum brasilense |
Substrates and Products (Substrate)
| EC Number | Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|---|
| 1.4.99.B4 | cis-4-hydroxy-D-proline + acceptor | the enzyme is involved in the metabolic pathway of trans-4-hydroxy-L-proline | Azospirillum brasilense | (4R)-DELTA1-pyrroline-4-hydroxy-2-carboxylate + reduced acceptor | - |
? | |
| 1.4.99.B4 | cis-4-hydroxy-D-proline + acceptor | the Vmax/Km value of the enzyme for cis-4-hydroxy-D-proline is about 124fold and about 736fold higher than the Vmax/Km values for trans-4-hydroxy-D-proline and D-proline, respectively. Both AbLhpBEF (heteromer of alpha-, beta- and gamma-subunit) and AbLhpB (catalytic beta-subunit) are strictly specific to cis-4-hydroxy-D-proline | Azospirillum brasilense | (4R)-DELTA1-pyrroline-4-hydroxy-2-carboxylate + reduced acceptor | - |
? | |
| 1.4.99.B4 | D-proline + acceptor | the Vmax/Km value of the enzyme for cis-4-hydroxy-D-proline is about 124fold and about 736fold higher than the Vmax/Km values for trans-4-hydroxy-D-proline and D-proline, respectively. Both AbLhpBEF (heteromer of alpha-, beta- and gamma-subunit) and AbLhpB (catalytic beta-subunit) are strictly specific to cis-4-hydroxy-D-proline | Azospirillum brasilense | ? | - |
? | |
| 1.4.99.B4 | trans-4-hydroxy-D-proline + acceptor | the Vmax/Km value of the enzyme for cis-4-hydroxy-D-proline is about 124fold and about 736fold higher than the Vmax/Km values for trans-4-hydroxy-D-proline and D-proline, respectively. Both AbLhpBEF (heteromer of alpha-, beta- and gamma-subunit) and AbLhpB (catalytic beta-subunit) are strictly specific to cis-4-hydroxy-D-proline | Azospirillum brasilense | ? | - |
? | |
| 4.2.1.77 | trans-3-hydroxy-L-proline | - |
Colwellia psychrerythraea | 1-pyrroline 2-carboxylate + H2O | - |
? | |
| 4.2.1.77 | trans-3-hydroxy-L-proline | - |
Azospirillum brasilense | 1-pyrroline 2-carboxylate + H2O | - |
? | |
| 4.2.1.77 | trans-3-hydroxy-L-proline | - |
Homo sapiens | 1-pyrroline 2-carboxylate + H2O | - |
? | |
| 5.1.1.8 | trans-4-hydroxy-L-proline | no activity with L-proline | Azospirillum brasilense | cis-4-hydroxy-D-proline | - |
? | |
Subunits
| EC Number | Subunits | Comment | Organism |
|---|---|---|---|
| 1.4.99.B4 | dodecamer | alpha4beta4gamma4, the gamma-subunit significantly contributes to stabilizing the alpha4beta4gamma4 structure (and the alpha-subunit), but not to enzyme catalysis | Azospirillum brasilense |
Synonyms
| EC Number | Synonyms | Comment | Organism |
|---|---|---|---|
| 1.4.99.B4 | AbLhpBEF | - |
Azospirillum brasilense |
| 1.4.99.B4 | C4DHyp dehydrogenase | - |
Azospirillum brasilense |
| 1.4.99.B4 | HYPDH | - |
Azospirillum brasilense |
| 1.4.99.B4 | LhpBEF | - |
Azospirillum brasilense |
| 4.2.1.77 | AbLhpH | - |
Azospirillum brasilense |
| 4.2.1.77 | CpLhpH | - |
Colwellia psychrerythraea |
| 4.2.1.77 | HsLhpH | - |
Homo sapiens |
| 4.2.1.77 | T3LHyp dehydratase | - |
Colwellia psychrerythraea |
| 4.2.1.77 | T3LHyp dehydratase | - |
Azospirillum brasilense |
| 4.2.1.77 | T3LHyp dehydratase | - |
Homo sapiens |
| 4.2.1.77 | T3LHypD | - |
Colwellia psychrerythraea |
| 4.2.1.77 | T3LHypD | - |
Azospirillum brasilense |
| 4.2.1.77 | T3LHypD | - |
Homo sapiens |
| 5.1.1.8 | AbLhpA | - |
Azospirillum brasilense |
| 5.1.1.8 | Hydroxyproline 2-epimerase | - |
Azospirillum brasilense |
| 5.1.1.8 | HypE | - |
Azospirillum brasilense |
Temperature Optimum [°C]
| EC Number | Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|---|
| 1.4.99.B4 | 30 | - |
assay at | Azospirillum brasilense |
| 4.2.1.77 | 30 | - |
assay at | Colwellia psychrerythraea |
| 4.2.1.77 | 30 | - |
assay at | Azospirillum brasilense |
| 4.2.1.77 | 30 | - |
assay at | Homo sapiens |
Temperature Stability [°C]
| EC Number | Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
|---|---|---|---|---|
| 1.4.99.B4 | 40 | - |
10 min, 70% loss of activity of the AbLhpB protein (catalytic subunit) | Azospirillum brasilense |
| 1.4.99.B4 | 50 | - |
10 min, the AbLhpBEF protein is stable, the AbLhpB protein (catalytic subunit) completely loses activity | Azospirillum brasilense |
pH Optimum
| EC Number | pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|---|
| 1.4.99.B4 | 9 | - |
assay at | Azospirillum brasilense |
Cofactor
| EC Number | Cofactor | Comment | Organism | Structure |
|---|---|---|---|---|
| 1.4.99.B4 | FAD | - |
Azospirillum brasilense | |
General Information
| EC Number | General Information | Comment | Organism |
|---|---|---|---|
| 1.4.99.B4 | metabolism | the enzyme is involved in the metabolic pathway of trans-4-hydroxy-L-proline | Azospirillum brasilense |
| 5.1.1.8 | metabolism | the enzyme is involved in the metabolic pathway of trans-4-hydroxy-L-proline | Azospirillum brasilense |
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