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Information on EC 1.5.5.2 - proline dehydrogenase and Organism(s) Escherichia coli and UniProt Accession P09546
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1.5.5.2 proline dehydrogenaseIUBMB Comments
A flavoprotein (FAD). The electrons from L-proline are transferred to the FAD cofactor, and from there to a quinone acceptor . In many organisms, ranging from bacteria to mammals, proline is oxidized to glutamate in a two-step process involving this enzyme and EC 1.2.1.88, L-glutamate gamma-semialdehyde dehydrogenase. Both activities are carried out by the same enzyme in enterobacteria.
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Word Map
- 1.5.5.2
-
putas
-
schizophrenia
- pyrroline-5-carboxylate
- delta1-pyrroline-5-carboxylate
-
flavoenzyme
-
psycho
-
hyperprolinemia
-
microdeletion
-
schizoaffective
-
2-acetyl-1-pyrroline
- rgs4
-
velocardiofacial
-
proline-dependent
-
digeorge
-
dysbindin
-
fragrant
- prolidase
- synthesis
The taxonomic range for the selected organisms is: Escherichia coli
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota, Archaea
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota, Archaea
Synonyms
prodh, proline dehydrogenase, prodh/pox, prodh1, l-proline dehydrogenase, dye-linked l-proline dehydrogenase, puta flavoprotein, ttprodh, jcprodh, tcprodh, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
proline dehydrogenase
bifunctional enzyme: proline utilization A (PutA)
proline/P5C dehydrogenase
bifunctional enzyme: proline dehydrogenase/L-glutamate gamma-semialdehyde dehydrogenase
PutA
L-proline dehydrogenase
-
-
-
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PRODH
proline oxidase
-
-
-
-
PutA flavoprotein
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combines DNA-binding, proline dehydrogenase and DELTA1-pyrroline-5-carboxylate dehydrogenase activities
PutA
bifunctional enzyme: proline dehydrogenase/L-glutamate gamma-semialdehyde dehydrogenase
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
L-proline + a quinone = (S)-1-pyrroline-5-carboxylate + a quinol
switching of enzyme from a cytosolic DNA-binding protein to a membrane-bound enzyme involves W211 in a flexible domain near the proline dehydrogenase active site and occurs in a time scale 10fold lower than the turnover number
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
redox reaction
-
-
-
-
oxidation
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-
-
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reduction
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-
-
-
PATHWAY SOURCE
PATHWAYS
-
-, -, -, -, -
SYSTEMATIC NAME
IUBMB Comments
L-proline:quinone oxidoreductase
A flavoprotein (FAD). The electrons from L-proline are transferred to the FAD cofactor, and from there to a quinone acceptor [3]. In many organisms, ranging from bacteria to mammals, proline is oxidized to glutamate in a two-step process involving this enzyme and EC 1.2.1.88, L-glutamate gamma-semialdehyde dehydrogenase. Both activities are carried out by the same enzyme in enterobacteria.
CAS REGISTRY NUMBER
COMMENTARY
9050-70-8
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SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
L-proline + acceptor
(S)-1-pyrroline-5-carboxylate + reduced acceptor
the bifunctional enzyme catalyzes the oxidation of proline in two steps. (S)-1-Pyrroline-5-carboxylate, the product of the first reaction is in spontaneous equilibrium with its tautomer L-glutamate gamma-semialdehyde: (1) L-proline + acceptor = (S)-1-pyrroline-5-carboxylate + reduced acceptor, (2) L-glutamate 5-semialdehyde + NAD+ + H2O = L-glutamate + NADH + H+
-
-
?
L-proline + oxidized dichlorophenolindophenol
1-pyrroline-5-carboxylate + reduced dichlorophenolindophenol
-
-
-
?
trans-4-hydroxy-L-proline + oxidized dichlorophenolindophenol
?
-
-
-
?
L-proline + 2,6-dichlorophenolindophenol + H2O
(S)-1-pyrroline-5-carboxylate + reduced 2,6-dichlorophenolindophenol
-
+ phenazine methosulfate as mediator
-
-
?
L-proline + 2,6-dichlorophenolindophenol + H2O
DELTA1-pyrroline-5-carboxylate + reduced 2,6-dichlorophenolindophenol
-
-
-
-
?
L-thiazolidine-4-carboxylate + acceptor + H2O
N-formylcysteine + reduced acceptor
-
-
-
?
L-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
PutA flavoprotein plays multiple roles in proline catabolism by functioning as a membrane-associated bi-functional enzyme and a transcriptional repressor of proline utilization genes
-
-
?
L-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
the enzyme catalyzes the first step of proline catabolism
-
-
?
L-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
-
-
-
?
L-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
-
-
-
?
L-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
-
-
-
?
L-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
-
-
-
-
?
L-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
-
enzyme exists in soluble and in membrane associated forms differing in catalytic properties
-
?
L-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
-
acceptors: 2,6-dichlorophenolindophenol, phenazine methosulfate, ferricyanide, menadione, cytochrome c
-
?
L-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
-
acceptors: 2,6-dichlorophenolindophenol, phenazine methosulfate, ferricyanide, menadione, cytochrome c
-
?
L-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
-
acceptors: 2,6-dichlorophenolindophenol, phenazine methosulfate, ferricyanide, menadione, cytochrome c
-
?
additional information
?
-
kinetic model for the overall PRODH-P5CDH reaction of bifunctional PutA enzyme. The intermediate is not released into the bulk medium, but the mechanism follows substrate channeling. The rate of NADH formation is 20fold slower than the steady-state turnover number for the overall reaction, The limiting rate constant observed for NADH formation in the first turnover increases by almost 40fold after multiple turnovers, achieving half of the steady-state value after 15 turnovers
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-
?
additional information
?
-
-
kinetic model for the overall PRODH-P5CDH reaction of bifunctional PutA enzyme. The intermediate is not released into the bulk medium, but the mechanism follows substrate channeling. The rate of NADH formation is 20fold slower than the steady-state turnover number for the overall reaction, The limiting rate constant observed for NADH formation in the first turnover increases by almost 40fold after multiple turnovers, achieving half of the steady-state value after 15 turnovers
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-
?
additional information
?
-
-
no substrate: D-proline, other L-amino acids, L-azetidine-2-carboxylate, succinate
-
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
L-proline + acceptor
(S)-1-pyrroline-5-carboxylate + reduced acceptor
the bifunctional enzyme catalyzes the oxidation of proline in two steps. (S)-1-Pyrroline-5-carboxylate, the product of the first reaction is in spontaneous equilibrium with its tautomer L-glutamate gamma-semialdehyde: (1) L-proline + acceptor = (S)-1-pyrroline-5-carboxylate + reduced acceptor, (2) L-glutamate 5-semialdehyde + NAD+ + H2O = L-glutamate + NADH + H+
-
-
?
L-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
PutA flavoprotein plays multiple roles in proline catabolism by functioning as a membrane-associated bi-functional enzyme and a transcriptional repressor of proline utilization genes
-
-
?
L-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
the enzyme catalyzes the first step of proline catabolism
-
-
?
L-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
-
-
-
?
L-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
-
-
-
?
L-proline + acceptor + H2O
(S)-1-pyrroline-5-carboxylate + reduced acceptor
-
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
FAD
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FAD may be released by 1 M KBr, apoprotein has no proline dehydrogenase activity but may be restored by external addition of FAD
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
N-propargylglycine
irreversibly inactivates PutA by covalently linking the flavin N(5) atom to the epsilon-amino of Lys329. Inactivation locks PutA into a conformation that may mimic the proline-reduced, membrane-associated form
5-ethylpentyl-barbituric acid
-
membrane-associated enzyme, not soluble enzyme
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
42
L-proline
-
in 50 mM potassium phosphate (pH 7.4, 50 mM NaCl), temperature not specified in the publication
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
5.2
L-proline
-
in 50 mM potassium phosphate (pH 7.4, 50 mM NaCl), temperature not specified in the publication
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
1.24
25°C, pH 7.5, proline oxidase activity of the bifunctional proline dehydrogenase/L-glutamate gamma-semialdehyde dehydrogenase
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
bifunctional proline dehydrogenase and delta1-pyrroline-5-carboxylate dehydrogenase, reactions of EC 1.5.5.2 and EC 1.2.1.88, respectively
UniProt
PutA669, corresponding to the N-terminal 669 residues of the 1320-amino acid PutA polypeptide chain
UniProt
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
137000
2 * 137000, bifunctional enzyme: proline dehydrogenase/L-glutamate gamma-semialdehyde dehydrogenase, SDS-PAGE
293000
gel filtration, bifunctional enzyme: proline dehydrogenase/L-glutamate gamma-semialdehyde dehydrogenase, sucrose density centrifugation
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
dimer
dimer
domain-swapped dimer with each subunit comprising three domains: a helical dimerization arm, a 120-residue domain containing a three-helix bundle similar to that in the helix-turn-helix superfamily of DNA-binding proteins and a beta/alpha-barrel PRODH domain with a bound lactate inhibitor
dimer
2 * 137000, bifunctional enzyme: proline dehydrogenase/L-glutamate gamma-semialdehyde dehydrogenase, SDS-PAGE
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
2.15 A resolution structure of the PRODH inactivated by N-propargylglycine. The initial events involved in broadcasting the reduced flavin state to the distal membrane-binding domain include major reorganization of the flavin ribityl chain, severe butterfly bending of the isoalloxazine ring, and disruption of an electrostatic network involving the flavin N(5) atom, Arg431, and Asp370. The active site is incompletely assembled in the absence of the substrate, and the binding of proline draws together conserved residues in helix 8 and the beta1-alphal loop to complete the active site
sitting-drop vaor diffusion method, structures of the PutA PRODH domain complexed with competitive inhibitors, acetate, L-lactate and L-tetrahydro-2-furoic acid
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
L432P
mutant form of PutA86-669, 5fold decrease in turnover-number and a severe loss in thermostability
D370A
-
mutant form of PutA86-601. The ratio of turnover-number to Km-value is 2.8fold lower than the ratio of the wild-type PutA86-601
D370A/Y540F
-
mutant form of PutA86-601. The ratio of turnover-number to Km-value is 15fold lower than the ratio of the wild-type PutA86-601
W194F
-
mutation in truncated enzyme containing residues 86-601, ratio of kcat to Km-value drops by 50%
W194F/W211F
-
mutation in truncated enzyme containing residues 86-601, ratio of kcat to Km-value drops by 95%
W211F
-
mutation in truncated enzyme containing residues 86-601, ratio of kcat to Km-value drops by 80%
Y540F
-
mutant form of PutA86-601. The ratio of turnover-number to Km-value is 5.8fold lower than the ratio of the wild-type PutA86-601
additional information
-
truncated enzyme containing residues 86-601 and only four Trp residues. Substantial conformational changes of truncated protein upon addition of proline
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
45
50% loss of activity of wild-type PutA86-669 after 1 h, 50% loss of of L432P mutant form of PutA86-669 after 8 min
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
gene product PutA functions as proline dehydrogenase and as repressor of genes putP and putA
-
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Wood, J.M.
Membrane association of proline dehydrogenase in Escherichia coli is redox dependent
Proc. Natl. Acad. Sci. USA
84
373-377
1987
Escherichia coli
Graham, S.B.; Stephenson, J.T.; Wood, J.M.
Proline dehydrogenase from Escherichia coli K12. Reconstitution of a functional membrane association
J. Biol. Chem.
259
2656-2661
1984
Escherichia coli
Scarpulla, R.C.; Soffer, R.L.
Membrane-bound proline dehydrogenase from Escherichia coli. Solubilization, purification, and characterization
J. Biol. Chem.
253
5997-6001
1978
Escherichia coli
Nadaraia, S.; Lee, Y.H.; Becker, D.F.; Tanner, J.J.
Crystallization and preliminary crystallographic analysis of the proline dehydrogenase domain of the multifunctional PutA flavoprotein from Escherichia coli
Acta Crystallogr. Sect. D
57
1925-1927
2001
Escherichia coli
Deutch, C.E.
Oxidation of L-thiazolidine-4-carboxylate by L-proline dehydrogenase in Escherichia coli
J. Gen. Microbiol.
138
1593-1598
1992
Escherichia coli
-
Brown, E.D.; Wood, J.M.
Conformational change and membrane association of the PutA protein are coincident with reduction of its FAD cofactor by proline
J. Biol. Chem.
268
8972-8979
1993
Escherichia coli
Zhang, M.; White, T.A.; Schuermann, J.P.; Baban, B.A.; Becker, D.F.; Tanner, J.J.
Structures of the Escherichia coli PutA proline dehydrogenase domain in complex with competitive inhibitors
Biochemistry
43
12539-12548
2004
Escherichia coli (P09546), Escherichia coli
Baban, B.A.; Vinod, M.P.; Tanner, J.J.; Becker, D.F.
Probing a hydrogen bond pair and the FAD redox properties in the proline dehydrogenase domain of Escherichia coli PutA
Biochim. Biophys. Acta
1701
49-59
2004
Escherichia coli
Lee, Y.H.; Nadaraia, S.; Gu, D.; Becker, D.F.; Tanner, J.J.
Structure of the proline dehydrogenase domain of the multifunctional PutA flavoprotein
Nat. Struct. Biol.
10
109-114
2003
Escherichia coli (P09546), Escherichia coli
Zhu, W.; Becker, D.F.
Exploring the proline-dependent conformational change in the multifunctional PutA flavoprotein by tryptophan fluorescence spectroscopy
Biochemistry
44
12297-12306
2005
Escherichia coli
Ostrander, E.L.; Larson, J.D.; Schuermann, J.P.; Tanner, J.J.
A conserved active site tyrosine residue of proline dehydrogenase helps enforce the preference for proline over hydroxyproline as the substrate
Biochemistry
48
951-959
2009
Escherichia coli (P09546), Escherichia coli
Srivastava, D.; Zhu, W.; Johnson, W.H.; Whitman, C.P.; Becker, D.F.; Tanner, J.J.
The structure of the proline utilization a proline dehydrogenase domain inactivated by N-propargylglycine provides insight into conformational changes induced by substrate binding and flavin reduction
Biochemistry
49
560-569
2010
Escherichia coli (P09546), Escherichia coli
Brown, E.D.; Wood, J.M.
Redesigned purification yields a fully functional PutA protein dimer from Escherichia coli
J. Biol. Chem.
267
13086-13092
1992
Escherichia coli (P09546), Escherichia coli
Moxley, M.A.; Becker, D.F.
Rapid reaction kinetics of proline dehydrogenase in the multifunctional proline utilization A protein
Biochemistry
51
511-520
2012
Escherichia coli
Moxley, M.A.; Sanyal, N.; Krishnan, N.; Tanner, J.J.; Becker, D.F.
Evidence for hysteretic substrate channeling in the proline dehydrogenase and delta1-pyrroline-5-carboxylate dehydrogenase coupled reaction of proline utilization A (PutA)
J. Biol. Chem.
289
3639-3651
2014
Escherichia coli (P09546), Escherichia coli
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