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Literature summary extracted from

  • Huijbers, M.M.; Martinez-Julvez, M.; Westphal, A.H.; Delgado-Arciniega, E.; Medina, M.; van Berkel, W.J.
    Proline dehydrogenase from Thermus thermophilus does not discriminate between FAD and FMN as cofactor (2017), Sci. Rep., 7, 43880 .
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.5.5.2 recombinant expression of the holo form of MBP-tagged TtProDH in Escherichia coli TOP10 cells, the recombinant enzyme contains about three times more FMN than FAD. The apoenzyme is produced by riboflavin auxotrophic Escherichia coli strain BSV11 and can be successfully reconstituted with FAD or FMN. The recombinant apo-enzyme reconstituted with FAD or FMN shows equal specific activities as holo-enzyme Thermus thermophilus

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
1.5.5.2 purified TtProDH mutant variant DELTAABC lacking helices alphaA, alphaB and alphaC, X-ray diffraction structure determination and analysis at 2.2 A resolution Thermus thermophilus

Protein Variants

EC Number Protein Variants Comment Organism
1.5.5.2 F10E/L12E construction of a variant with a more polar N-terminus, mutant F10E/L12E, because Thermus thermophilus ProDH (TtProDH), produced through fusion with maltose-binding protein (MBP) appears to be prone to aggregation. Preparation of TtProDH mutant variant DELTAABC, which lacks helices alphaA, alphaB and alphaC, the mutant shows no electron density for an AMP moiety of the cofactor Thermus thermophilus

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.5.5.2 36.6
-
 L-proline recombinant apo-enzyme reconstituted with FAD, pH 7.4, 25°C, with 2,6-dichlorphenol-indophenol Thermus thermophilus
1.5.5.2 50.1
-
 L-proline recombinant apo-enzyme reconstituted with FMN, pH 7.4, 25°C, with 2,6-dichlorphenol-indophenol Thermus thermophilus
1.5.5.2 76.6
-
 L-proline recombinant holo-enzyme, pH 7.4, 25°C, with 2,6-dichlorphenol-indophenol Thermus thermophilus

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.5.5.2 L-proline + a quinone Thermus thermophilus the reverse reaction is catalyzed by pyrroline-5-carboxylate reductase, EC 1.5.1.2 (S)-1-pyrroline-5-carboxylate + a quinol
-
 ir
1.5.5.2 L-proline + a quinone Thermus thermophilus HB27 / ATCC BAA-163 / DSM 7039 the reverse reaction is catalyzed by pyrroline-5-carboxylate reductase, EC 1.5.1.2 (S)-1-pyrroline-5-carboxylate + a quinol
-
 ir

Organism

EC Number Organism UniProt Comment Textmining
1.5.5.2 Thermus thermophilus Q72IB8
-
-
1.5.5.2 Thermus thermophilus HB27 / ATCC BAA-163 / DSM 7039 Q72IB8
-
-

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
1.5.5.2 0.84
-
 purified recombinant apo-enzyme, pH 7.4, 25°C, with 2,6-dichlorphenol-indophenol Thermus thermophilus
1.5.5.2 4.74
-
 purified recombinant holo-enzyme, pH 7.4, 25°C, with 2,6-dichlorphenol-indophenol Thermus thermophilus
1.5.5.2 4.83
-
 purified recombinant apo-enzyme reconstituted with FAD, pH 7.4, 25°C, with 2,6-dichlorphenol-indophenol Thermus thermophilus
1.5.5.2 4.95
-
 purified recombinant apo-enzyme reconstituted with FMN, pH 7.4, 25°C, with 2,6-dichlorphenol-indophenol Thermus thermophilus

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.5.5.2 L-proline + 2,6-dichlorphenol-indophenol
-
 Thermus thermophilus (S)-1-pyrroline-5-carboxylate + reduced 2,6-dichlorphenol-indophenol
-
 ?
1.5.5.2 L-proline + 2,6-dichlorphenol-indophenol
-
 Thermus thermophilus HB27 / ATCC BAA-163 / DSM 7039 (S)-1-pyrroline-5-carboxylate + reduced 2,6-dichlorphenol-indophenol
-
 ?
1.5.5.2 L-proline + a quinone the reverse reaction is catalyzed by pyrroline-5-carboxylate reductase, EC 1.5.1.2 Thermus thermophilus (S)-1-pyrroline-5-carboxylate + a quinol
-
 ir
1.5.5.2 L-proline + a quinone the reverse reaction is catalyzed by pyrroline-5-carboxylate reductase, EC 1.5.1.2 Thermus thermophilus HB27 / ATCC BAA-163 / DSM 7039 (S)-1-pyrroline-5-carboxylate + a quinol
-
 ir

Subunits

EC Number Subunits Comment Organism
1.5.5.2 dimer 2 * 74401, recombinant MBP-tagged enzyme, sequence calculation, 2 * 71899, recombinant enzyme mutant DELTAABC, sequence calculation Thermus thermophilus

Synonyms

EC Number Synonyms Comment Organism
1.5.5.2 TtProDH
-
 Thermus thermophilus

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.5.5.2 25
-
 assay at Thermus thermophilus

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.5.5.2 7.2
-
 L-proline recombinant apo-enzyme reconstituted with FAD, pH 7.4, 25°C, with 2,6-dichlorphenol-indophenol Thermus thermophilus
1.5.5.2 7.9
-
 L-proline recombinant apo-enzyme reconstituted with FMN, pH 7.4, 25°C, with 2,6-dichlorphenol-indophenol Thermus thermophilus
1.5.5.2 9.8
-
 L-proline recombinant holo-enzyme, pH 7.4, 25°C, with 2,6-dichlorphenol-indophenol Thermus thermophilus

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.5.5.2 7.4
-
 assay at Thermus thermophilus

Cofactor

EC Number Cofactor Comment Organism Structure
1.5.5.2 FAD proline dehydrogenase from Thermus thermophilus does not discriminate between FAD and FMN as cofactor, redox active prosthetic group Thermus thermophilus
1.5.5.2 FMN proline dehydrogenase from Thermus thermophilus does not discriminate between FAD and FMN as cofactor, redox active prosthetic group Thermus thermophilus
1.5.5.2 additional information the recombinant holo form of MBP-tagged TtProDH, as produced in Escherichia coli TOP10 cells, contains about three times more FMN than FAD. The crystal structure of TtProDH variant DELTAABC, which lacks helices alphaA, alphaB and alphaC, shows no electron density for an AMP moiety of the cofactor. ProDH adopts a distorted (betalpha)8 TIM-barrel fold and is the only known TIM-barrel enzyme that contains an FAD cofactor. One cofactor molecule per enzyme subunit Thermus thermophilus

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
1.5.5.2 146
-
 L-proline recombinant holo-enzyme, pH 7.4, 25°C, with 2,6-dichlorphenol-indophenol Thermus thermophilus
1.5.5.2 158
-
 L-proline recombinant apo-enzyme reconstituted with FMN, pH 7.4, 25°C, with 2,6-dichlorphenol-indophenol Thermus thermophilus
1.5.5.2 198
-
 L-proline recombinant apo-enzyme reconstituted with FAD, pH 7.4, 25°C, with 2,6-dichlorphenol-indophenol Thermus thermophilus