Information on EC 2.7.9.1 - pyruvate, phosphate dikinase and Organism(s) Zea mays and UniProt Accession P11155

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Zea mays
UNIPROT: P11155
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The expected taxonomic range for this enzyme is: Bacteria, Eukaryota, Archaea


The taxonomic range for the selected organisms is: Zea mays

EC NUMBER
COMMENTARY hide
2.7.9.1
-
RECOMMENDED NAME
GeneOntology No.
pyruvate, phosphate dikinase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + pyruvate + phosphate = AMP + phosphoenolpyruvate + diphosphate
show the reaction diagram
sequential mechanism for the addition of ATP and phosphate and a ping-pong mechanism for the addition of pyruvate and release of phosphoenolpyruvate
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
phospho group transfer
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-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
C4 photosynthetic carbon assimilation cycle, NAD-ME type
-
-
C4 photosynthetic carbon assimilation cycle, NADP-ME type
-
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C4 photosynthetic carbon assimilation cycle, PEPCK type
-
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L-glutamine biosynthesis III
-
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C4 and CAM-carbon fixation
-
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photosynthesis
-
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Pyruvate metabolism
-
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Carbon fixation in photosynthetic organisms
-
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Carbon fixation pathways in prokaryotes
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Metabolic pathways
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Microbial metabolism in diverse environments
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-
SYSTEMATIC NAME
IUBMB Comments
ATP:pyruvate, phosphate phosphotransferase
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CAS REGISTRY NUMBER
COMMENTARY hide
9027-40-1
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
physiological function
PPDK is the key enzyme of the C4 pathway, and its activity may limit the photosynthesis rate in maize leaves. The amount of PPDK (unphosphorylated) involved in C4 photosynthesis is strictly controlled by light intensity. Diverse regulatory pathways may work alone or in combination to fine-tune C4PPDK activity in response to changes in light
evolution
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three-dimensional modeling of PPDKs from divergent organisms and comparion of the orientation of the phosphorylatable histidine residue within the central domain of PPDKs. These PPDKs are compared using a maximum-likelihood tree. Phylogenetic analysis of the N- and C-terminal sequences of PPDKs from different species, overview
metabolism
physiological function
additional information
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role of the N- and C-termini on the orientation of the PPDK central domain, three-dimensional structure analysis
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ATP + pyruvate + phosphate
AMP + phosphoenolpyruvate + diphosphate
show the reaction diagram
AMP + phosphoenolpyruvate + diphosphate
ATP + pyruvate + phosphate
show the reaction diagram
ATP + pyruvate + arsenate
?
show the reaction diagram
-
-
-
-
ir
ATP + pyruvate + phosphate
AMP + phosphoenolpyruvate + diphosphate
show the reaction diagram
additional information
?
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NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
ATP + pyruvate + phosphate
AMP + phosphoenolpyruvate + diphosphate
show the reaction diagram
AMP + phosphoenolpyruvate + diphosphate
ATP + pyruvate + phosphate
show the reaction diagram
ATP + pyruvate + phosphate
AMP + phosphoenolpyruvate + diphosphate
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Mg2+
required
K+
-
more than 20fold activation pH 7.4 and at pH 8.1
Mn2+
-
not activation
Na+
-
slight activation of the phosphoenol pyruvate formation
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
ilimaquinone
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uncompetitive/mixed type versus pyruvate and versus ATP, 48% inhibition of C4 acid cycle evolution. IC50: 0.292 mM
ATP
-
-
diphosphate
gamma(p-Arsenophenyl)-n-butyrate
-
-
ilimaquinone
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selectively toxic to C4 plants, isolated from a marine sponge, heterocyclic compound, 3 rings, 2-D structure shown
MgHPO4
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competitive to HPO42-
p-chloromercuribenzoate
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p-hydroxymercuribenzoate
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-
phosphate
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phosphoenolpyruvate
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competitive to pyruvate
PPDK regulatory protein
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RP catalyzes reversible phosphorylation on T456 by RP, inactive form phoshorylated at T456, ADP-dependent, a catalytic His-phophorylation precedes phosphorylation at T456, pyruvate (2 mM) can inhibit inactivation by RP
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pyruvate
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unguinol
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from fungal isolate F3000054, a mixed noncompetitive inhibitor of PPDK with respect to the substrates pyruvate and ATP and an uncompetitive inhibitor of PPDK with respect to phosphate. Unguinol has deleterious effects on a model C4 plant but no effect on a model C3 plant, effects in vivo, overview; mixed noncompetitive inhibition of PPDK with respect to the substrates pyruvate and ATP, uncompetitive with respect to phosphate, phytotoxic effects (bleaching) on C4 plants, no effect on a model C3 plant (Hordeum vulgare)
additional information
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
thiols
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required for activity in solution
additional information
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.009 - 0.095
ATP
0.04
diphosphate
0.134 - 1.5
phosphate
0.046 - 0.194
phosphoenolpyruvate
0.065 - 0.25
pyruvate
additional information
additional information
-
kinetics at different temperature, the t1/2 for dephosphorylation of PPDK from chilling-acclimated Miscanthus x giganteus leaves increased to 3.1fold compared to warm-grown leaves, overview
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Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.13
AMP
-
pH 8.1, 25ºC, phosphoenolpyruvate formation
0.036
ATP
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pH 8, 22ºC
0.32
diphosphate
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pH 8.1, 25ºC, phosphoenolpyruvate formation
0.15
phosphoenolpyruvate
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pH 8.1, 25ºC, phosphoenolpyruvate formation
additional information
additional information
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inhibition kinetics for unguinol on PPDK
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IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.292
ilimaquinone
Zea mays;
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uncompetitive/mixed type versus pyruvate and versus ATP, 48% inhibition of C4 acid cycle evolution. IC50: 0.292 mM
0.29
ilimaquinone
Zea mays;
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-
0.04
unguinol
Zea mays;
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.043
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T456F mutant
0.067
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inactive enzyme obtained from dark-grown plants
0.1 - 0.19
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leaf enzyme extract, pH and temperature not specified in the publication
0.24
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T456Y mutant
1.2
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measured at 30ºC
2.91
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after activation by phosphate of an enzyme obtained from dark-grown plants
4.1
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4.5
-
-
additional information
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specific activity of PPDK in plants transformed with the construct showing 17 amino acid substitutions inversely correlates with amount of enzyme in leaves, PPDK activity becomes constant when the amount is above 5 mg/g fresh weight, enzyme contents estimated by Western blot, spectrometric assay at 25°C, two transformants retained 70% of activity after 180 min indicating a comparable cold-tolerance like Flaveria brownii plants, further effect of cold-tolerant PPDK on photosynthetic rate estimated by measurement of CO2 uptake at leaf temperatures of 30°C, 20°C, 13°C, and 8°C, increase by 23% at 8°C shown, no effects at higher temperatures
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
8
assay at
6.9
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pyruvate formation
7
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the pyruvate-forming reaction is strongly favoured at pH 7.0
8.2
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phosphoenolpyruvate formation
8.3
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the competency of the enzyme in catalyzing its phosphoenolpyruvate-forming reaction at pH 7.0 is dramatically reduced, having only 6% of the rate at pH 8.3
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7 - 8.3
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the competency of the enzyme in catalyzing its phosphoenolpyruvate-forming reaction at pH 7.0 is dramatically reduced, having only 6% of the rate at pH 8.3
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30
assay at
22
-
assay at
30
assay at
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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cold tolerance of recombinant maize plants estimated, crude leaf extracts placed on ice to measure residual PPDK activities, PPDK extracted from untransformed maize looses 90% of its activity after 20 min, a retained PPDK activity 70% of after 180 min in two transformants corresponds to a cold-tolerance shown for Flaveria brownii plants
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.2
isoelectric focusing (pH range from 4 to 7), cytosolic CyPPDK 2
5.3
isoelectric focusing (pH range from 4 to 7), cytosolic CyPPDK 2
5.4
isoelectric focusing (pH range from 4 to 7), cytosolic CyPPDK 2
additional information
annotation of different isoforms of one of the two cytosolic isoforms of PPDK of maize
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
time course analysis of seven developmental stages of maize, 4-40 days after pollination, significant PPDK accumulation at 21 days after pollination
Manually annotated by BRENDA team
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
additional information
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enzyme activity at different climates, cold climates at 14°C and warm climates at 25°C, overview
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
high levels of PPDK protein accumulate in mesophyll chloroplasts, identification of a transit peptide cleavage site, the mature isozyme C4PPDK contains the specific N-terminal sequence TTKK
Manually annotated by BRENDA team
seven isoforms of cytosolic PPDK of maize identified by proteomics, distinguished by deduced molecular mass, isoelectric point and slight differences in primary peptide sequence, in silico prediction of subcellular destination
Manually annotated by BRENDA team
the mature isozyme CyPPDKZm1 contains the 11-residue sequence MAPAPCGRSSQ
Manually annotated by BRENDA team
PDB
SCOP
CATH
UNIPROT
ORGANISM
Zea mays;
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
99280
cytosolic CyPPDK 2
104900
cytosolic CyPPDK 2
105100
cytosolic CyPPDK 2
105700
cytosolic CyPPDK 2
105000
D3K3U9;
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370000
387000
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sedimentation analysis
additional information
different isoforms of one of the two cytosolic isoforms of PPDK of maize distinguished by pI value, molecular mass and altered amino acid positions in identified peptides by isoelectric focusing, 2-D electrophoresis and liquid chromatography
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homotetramer
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4 * 95000, the enzyme is maximally active as a homotetramer and is inactive in the dimeric and monomeric forms
tetramer
additional information
-
role of the N- and C-termini on the orientation of the PPDK central domain, three-dimensional structure analysis
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
phosphoprotein
in C4 plants, pyruvate orthophosphate dikinase (PPDK) activity is tightly dark/light regulated by reversible phosphorylation of an active-site Thr residue, catalyzed by PPDK regulatory protein (PDRP). Phosphorylation and dephosphorylation of PPDK lead to its inactivation and activation, respectively. Light intensity rather than the light/dark transition regulates PPDK activity by modulating the reversible phosphorylation at Thr527 of PPDK in Zea mays. PDRP catalyzes the reversible phosphorylation of PPDK rapidly and efficiently, but a subtle conformational change around PPDK's active site Thr527 will affect its catalytic efficiency. Phosphorylation at Thr527 of PPDK is independent of the light/dark transition. The plastidic isozyme shows phosphorylation at four serine (Ser)/Thr residues, Thr527, Ser528, Thr309, and Ser506 are targets of PDRP. The two hydrogen bonds between the highly conserved residues Ser528 and Gly525 are required for PDRP-mediated phosphorylation of the active-site Thr527 of the isozyme. Analysis by high resolution mass spectrometry, locations of the four phosphorylation residues in the primary sequence and the three-dimensional structure of C4PPDK, modelling, overview
phosphoprotein
additional information
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
hanging-drop vapour diffusion method, crystal strcuture with and without phosphoenolpyruvate, determined at 2.3 A resolution
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method description
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
10
-
30 min, about 40% loss of activity
12
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loses activity below at about 12°C by dissociation of the tetramer, considered as one possible cause of the reduction of the photosynthetic rate of maize at low temperatures
20 - 40
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stable for at least 30 min
50
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irreversible denaturation
additional information
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
glycerol protects both the day-form and night-form in vitro
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Mg2+ stabilizes the oligomeric structure of the enzyme
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
0ºC, as a precipitate in a 66% saturated solution of (NH4)2SO4
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
native isozymes from leaves, subcellular fractionation, recombinant tagged wild-type and mutant enzymes from Escherichia coli strain BL21 (RIL) by immobilized metal affinity chromatography
isolation of an inactive enzyme form dark-grown leaves
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maize PPDK of leaves expressed for inhibitor studies in Escherichia coli with a His-tag and purified on a Ni-affinity column
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method that includes successive chromatography through DE-52, hydroxyapatite, Sephadex G-200 and Blue agarose
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native isozymes from leaves, subcellular fractionation
partial
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partial, using Sephadex-G200 and Hypatite C chromatography
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recombinant maize PPDK used for inactivation/activation assay by the PPDK regulatory protein RP
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Escherichia coli, three constructs, one construct consisting of the 3'-part of Flaveria brownii (Asteraceae) of cold tolerant PPDK fused to maize PPDK (15th exon), another construct includes a set of point mutations to substitute all of the 17 residues that differ between the 3'-parts of maize and Flaveria brownii PPDK, respectively, the whole genomic sequence of the maize PPDK gene is included as a control, transformation of constructs into maize inbred line A188 by Agrobacterium tumefaciens (strain LBA4404), individual range of variation in the amount of PPDK among regenerated plants, crude leaf extracts of some transformed plants produce a large amount of cold tolerant recombinant enzyme and reveal a greatly improved cold tolerance especially by using the construct altered at 17 amino acid positions
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gene C4ppdkZm1, quantitative real-time PCR enzyme expression analysis, recombinant expression of tagged wild-type and mutant enzymes in Escherichia coli strain BL21 (RIL)
; expressed in Escherichia coli
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expressed in Arabidopsis thaliana, found exclusively in chloroplasts of transgenic Arabidopsis plants
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expressed in Escherichia coli
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expressed in Oryza sativa subsp. indica cultivar IR64
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gene CyppdkZm2, quantitative real-time PCR enzyme expression analysis
gene PPDK, phylogenetic analysis of the N- and C-terminal sequences of PPDKs from different species, overview
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gene ppdk, Triticum aestivum cv. Zhoumai19 immature embryosare transfected by particle bombardment with chimeric genes containing pepc or ppdk cDNA and the bialaphos resistance gene under the control of the Cauliflower mosaic virus 35S promoter. Three types of transgenic lines containing 1. pepc cDNA (PC lines), 2. ppdk cDNA (PK lines), or 3. both genes (PKC lines) are obtained through callus differentiation, phosphinthricine resistance screening, plantlet regeneration, and molecular detection. Quantitative real-time PCR enzyme expression analysis. Net photosynthetic rates of mutants are increased compared to controls
D3K3U9;
quantitative expression analysis of the enzyme during endosperm development, expression profiles, overview
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quantitative real-time reverse transcription (RT)-PCR expression analysis during transfer of plants from 25°C to 14°C growth temperature
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to improve the cold stability of the enzyme, a cold-tolerant PPDK cDNA isolated from Flaveria brownii is introduced into maize by Agrobacterium-mediated transformation. Higher levels of expression are ontained by using a double intron cassette and a chimeric cDNA made from Flaveria bidentis and Flaveria brownii with a maximum content of 1 mg/g fresh weight. In leaves of transgenic maize, PPDK molecules produced from the transgene are detected in cold-tolerant homotetramers or in heterotetramers of intermediate cold susceptibility formed with the internal PPDK. A significant improvement in the cold stability of PPDK can be achieved when a suffcient quantity of cold-tolerant subunits is expressed in transgenic maize leaves
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
in the C4 pathway, enzyme activity is strictly regulated in an up/down manner by the level of incident light
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
G525A
site-directed mutagenesis, the mutant shows no phosphorylation signal
G525P
site-directed mutagenesis, the mutant shows no phosphorylation signal
H529A
site-directed mutagenesis
S506A
site-directed mutagenesis
S528A
site-directed mutagenesis
S528C
site-directed mutagenesis, the mutant shows a phosphorylation signal only slightly weaker than the wild-type enzyme due to the tighter binding of S528 to G525 compared to C528
S528D
site-directed mutagenesis
S528T
site-directed mutagenesis, the mutant shows no phosphorylation signal
S528Y
site-directed mutagenesis, the mutant shows no phosphorylation signal
T309A
site-directed mutagenesis
T527A
site-directed mutagenesis
T527D
site-directed mutagenesis
H458N
-
no activity
T456D
-
no activity
T456E
-
no activity
T456S
-
111% activity with respect to wild type
T456V
-
98% activity with respect to wild type
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
agriculture
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to improve the cold stability of the PPDK enzyme, studies on cold tolerance in plants
agriculture
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modification of the cold sensitivity of a C4 photosynthetic enzyme by employing cereal transformation technology. A significant improvement in the cold stability of PPDK can be achieved when a suffcient quantity of cold-tolerant subunits is expressed in transgenic maize leaves
drug development
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the enzyme is a potential herbicide target in C4 plants