Information on EC 3.5.4.16 - GTP cyclohydrolase I

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The expected taxonomic range for this enzyme is: Eukaryota, Bacteria, Archaea

EC NUMBER
COMMENTARY hide
3.5.4.16
-
RECOMMENDED NAME
GeneOntology No.
GTP cyclohydrolase I
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
GTP + H2O = formate + 7,8-dihydroneopterin 3'-triphosphate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
amidine hydrolysis
-
-
-
-
C-N bond cleavage
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
6-hydroxymethyl-dihydropterin diphosphate biosynthesis I
-
-
drosopterin and aurodrosopterin biosynthesis
-
-
Folate biosynthesis
-
-
Metabolic pathways
-
-
preQ0 biosynthesis
-
-
sulfopterin metabolism
-
-
tetrahydrobiopterin biosynthesis I
-
-
tetrahydrobiopterin biosynthesis II
-
-
tetrahydrobiopterin biosynthesis III
-
-
tetrahydrofolate metabolism
-
-
tetrahydromonapterin biosynthesis
-
-
SYSTEMATIC NAME
IUBMB Comments
GTP 7,8-8,9-dihydrolase
The reaction involves hydrolysis of two C-N bonds and isomerization of the pentose unit; the recyclization may be non-enzymic. This enzyme is involved in the de novo synthesis of tetrahydrobiopterin from GTP, with the other enzymes involved being EC 1.1.1.153 (sepiapterin reductase) and EC 4.2.3.12 (6-pyruvoyltetrahydropterin synthase) [3].
CAS REGISTRY NUMBER
COMMENTARY hide
37289-19-3
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
Q8S3C2
SwissProt
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
strain MJ0775
-
-
Manually annotated by BRENDA team
strain MJ0775
-
-
Manually annotated by BRENDA team
Mus musculus C57BL/6
strain C57BL/6
-
-
Manually annotated by BRENDA team
Mus musculus C57BL6
-
-
-
Manually annotated by BRENDA team
strain NRRL 5646
SwissProt
Manually annotated by BRENDA team
Nocardia sp. NRRL 5646
strain NRRL 5646
SwissProt
Manually annotated by BRENDA team
swallowtail butterfly
-
-
Manually annotated by BRENDA team
-
O9701
UniProt
Manually annotated by BRENDA team
Serratia indica
-
SwissProt
Manually annotated by BRENDA team
spinach
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
strain HB8
Uniprot
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
metabolism
physiological function
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
2-amino-5-formylamino-6-ribofuranosylamino-4(3H)-pyrimidinone triphosphate + H2O
formate + 2-amino-4-hydroxy-6-(erythro-1,2,3-trihydroxypropyl)-dihydropteridine triphosphate
show the reaction diagram
substrate is the reaction intermediate of the overall reaction
-
-
?
beta,gamma-methyleneguanosine 5'-triphosphate + H2O
dihydroneopterin 2',3'-cyclic phosphate + ?
show the reaction diagram
beta-gamma-methyleneguanosine 5'-triphosphate + H2O
beta-gamma-methylene-7,8-dihydroneopterin 3'-triphosphate + formate
show the reaction diagram
-
-
-
?
GDP + H2O
dihydroneopterin 2',3'-cyclic phosphate + ?
show the reaction diagram
GDP + H2O
formate + 2-amino-4-hydroxy-6-(erythro-1,2,3-trihydroxypropyl)-dihydropteridine diphosphate
show the reaction diagram
-
i.e. dihydroneopterin 3'-triphosphate
-
?
GTP + H2O
2-amino-5-formylamino-6-ribosylamino-4(3H)-pyrimidinone + ?
show the reaction diagram
-
mutant H179N is not able to perform the whole reaction step
-
-
r
GTP + H2O
dihydroneopterin triphosphate + formate
show the reaction diagram
GTP + H2O
formate + 2-amino-4-hydroxy-6-(erythro-1,2,3-trihydroxypropyl)-dihydropteridine triphosphate
show the reaction diagram
GTP + H2O
formate + 2-amino-4-hydroxy-6-(erythro-1,2,3-trihydroxypropyl)dihydropteridine triphosphate
show the reaction diagram
GTP + H2O
formate + 7,8-dihydro-D-neopterin 2',3'-cyclic phosphate + diphosphate
show the reaction diagram
-
-
-
-
?
GTP + H2O
formate + 7,8-dihydroneopterin 3'-triphosphate
show the reaction diagram
GTP + H2O
formate + D-erythro-dihydroneopterin triphosphate
show the reaction diagram
guanosine 5'-[gamma-thio]triphosphate + H2O
dihydroneopterin 2',3'-cyclic phosphate + ?
show the reaction diagram
-
-
-
-
?
additional information
?
-
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
GTP + H2O
dihydroneopterin triphosphate + formate
show the reaction diagram
GTP + H2O
formate + 2-amino-4-hydroxy-6-(erythro-1,2,3-trihydroxypropyl)-dihydropteridine triphosphate
show the reaction diagram
GTP + H2O
formate + 2-amino-4-hydroxy-6-(erythro-1,2,3-trihydroxypropyl)dihydropteridine triphosphate
show the reaction diagram
GTP + H2O
formate + 7,8-dihydroneopterin 3'-triphosphate
show the reaction diagram
GTP + H2O
formate + D-erythro-dihydroneopterin triphosphate
show the reaction diagram
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Fe2+
-
dependent, contains 1 Fe2+ per protomer
Li+
Serratia indica
-
activates
Na+
Serratia indica
-
activates
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
(6R)-L-erythro-5,6,7,8-tetrahydrobiopterin
2,4-Diamino-6-hydroxypyrimidine
7-methyl-GTP
-
36% inhibition at 2 mM GMP
8-Aminoguanosine triphosphate
-
-
8-Azaguanine
-
43% inhibition at 2.9 mM
8-ethoxycarbonyl-7-deazaguanine
-
78% inhibition at 2.9 mM
8-methyl-7-deazaguanine
-
94% inhibition at 2.9 mM
8-oxo-dGTP
competitive inhibition
8-oxo-GTP
8-trifluoromethyl-7-deazaguanine
-
17% inhibition at 2.9 mM
ADP
-
competitive inhibitor
Al3+
Serratia indica
-
-
ascorbate
-
-
caveolin-1
-
overexpression of caveolin-1 inhibits GTPCH I activity
-
Co3+
-
no activity detected regardless of metal concentration
DAHP
-
non-competitive inhibitor
dihydrobiopterin
-
induction of feedback inhibition, binding site structure
Divalent cations
-
-
-
EDTA
-
no activity of GCYH-IB is observed in the presence of EDTA
fapy-GMP
-
1 mM results in a 5fold reduction in activity
GTP cyclohydrolase feedback regulatory protein
-
i.e. GFRP, GTP cyclohydrolase I, GTPCH-1, undergoes negative feedback regulation by its endproduct tetrahydrobiopterin via interaction with the GTP cyclohydrolase feedback regulatory protein, GFRP. GFRP binding increased the apparent Km of GTPCH-1, which also contributes to inhibition and increases the cooperativity of substrate binding in the wild-type but not the S81D mutant. GFRP both inhibits and stimulates GTPCH-1 activity in vitro depending on interactions with either tetrahydrobiopterin or phenylalanine. GTPCH-1 phosphorylation reduces its binding to GFRP
-
GTP cyclohydrolase I feedback regulatory protein GFRP
-
natural inhibitor, inhibition mechanism, in vitro inhibition together with 2,4-diamino-6-hydroxypyrimidine, the inhibition is fully reversible by L-phenylalanine, regulatory function in physiological feedback inhibition, overview
-
GTPCH feedback regulatory protein
-
GFRP, the allosteric regulatory protein GFRP triggers a noncompetitive attenuation of GTPCH activity, an allosteric effector is unnecessary for GFRP to influence GTPCH activity. GFRP-mediated allosteric regulation by small molecule effectors is indistinguishable for truncated mutant DELTA45-GTPCH and wild-type GTPCH
-
guanosine 5'-tetraphosphate
-
-
H2O2
-
more than 0.3 mM H2O2 result in a decrease in activity of GTPCHI, the function of the GTP cyclohydrolase I/GTP cyclohydrolase I feedback regulatory protein complex is not affected by H2O2
L-erythro-5,6,7,8-tetrahydrobiopterin
L-erythro-7,8-dihydrobiopterin
-
-
L-Sepiapterin
-
-
N-acetyl-serotonin
p-chloromercuribenzoate
Serratia indica
-
-
SO42-
Serratia indica
-
-
streptozotocin
-
the expression of GCH-I is decreased by streptozotocin treatment (60 mg/kg iv, 7 weeks)
tetrahydrobiopterin
TTP
-
competitive inhibitor
additional information
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
17beta-estradiol
-
can regulate GTPCH gene expression via transcriptional mechanisms
H2O2
-
0.1 mM H2O2 is the optimum concentration for the activation of GTPCHI
interferon-gamma
-
GCH1 expression is strongly induced by a mixture of interleukin-1beta, tissue necrosis factor-alpha, and interferon-gamma released by microglia under brain-damaging conditions
-
Interleukin-1beta
-
GCH1 expression is strongly induced by a mixture of interleukin-1beta, tissue necrosis factor-alpha, and interferon-gamma released by microglia under brain-damaging conditions
-
L-phenylalanine
lipolysaccharide
commercially isolated from Escherichia coli serotype 026:B6, activates and upregulates the enzyme in adipose tissue, but in cultured 3T3-L1 adipocytes lipolysaccharide addition only increases the enzyme expression level in presence of TNFalpha and interferon gamma, overview
-
tissue necrosis factor-alpha
-
GCH1 expression is strongly induced by a mixture of interleukin-1beta, tissue necrosis factor-alpha, and interferon-gamma released by microglia under brain-damaging conditions
-
additional information
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0143
GDP
pH 8.5, 25C, recombinant enzyme
0.004 - 980
GTP
additional information
additional information
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0017
GDP
Thermus thermophilus
Q5SH52
pH 8.5, 25C, recombinant enzyme
0.0011 - 0.039
GTP
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.00022
8-oxo-dGTP
pH 8.5, 25C, recombinant enzyme
0.0000054
8-oxo-GTP
pH 8.5, 25C, recombinant enzyme
0.071
ATP
pH 8.5, 25C, recombinant enzyme
0.02
dGTP
pH 8.5, 25C, recombinant enzyme
0.173
GTP
pH 8.0, 37C, recombinant enzyme
additional information
additional information
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.000001
wild-type native enzyme and recombinant HAT10-tagged enzyme from expression in Escherichia coli BL21(DE3)
0.000029
recombinant enzyme from expression in Mycobacterium smegmatis/pSMT-260
0.00027
recombinant HAT-tagged enzyme from expression in Escherichia coli JM109
0.00088
recombinant HAT-217-tagged enzyme from expression in Escherichia coli BL21(DE3)
0.088
wild-type enzyme, substrate 2-amino-5-formylamino-6-ribofuranosylamino-4(3H)-pyrimidinone triphosphate
0.091
wild-type enzyme, substrate GTP
6
purified recombinant HAT-tagged enzyme expressed in Escherichia coli
67
Serratia indica
-
-
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7 - 8.5
-
mutant V150E
7
-
mutant S135C
7.3
-
phosphate buffer: 2 optima, pH 7.3 and 8.0
8 - 8.5
-
-
8.6
Serratia indica
-
-
additional information
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7 - 11
O9701
-
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
26
-
assay at
45
-
mutant E152K
50
O9701
the enzyme activity is improved by the rise in temperature up to 50C
55
-
mutants S135C and R185G
65
-
mutants H112D, K136E, and H179Q
70
-
mutant H113 N
72
-
mutant R56L
75
-
mutants C110G and L134Q
85
-
mutants E111K and V150E
90
-
mutant C181S
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
38 - 78
-
half-maximal activity at
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
adipocyte cell line
Manually annotated by BRENDA team
GCHI mRNA level is highest in unripe fruit
Manually annotated by BRENDA team
-
the black markings on the larva during fourth ecdysis are involving the GTPCHI activity
Manually annotated by BRENDA team
-
transfected with a tet-off plasmid and a plasmid encoding human GCH1, the stable clone is named tet-GCH cell
Manually annotated by BRENDA team
additional information
GCH expression patterns
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Listeria monocytogenes serovar 1/2a (strain ATCC BAA-679 / EGD-e)
Neisseria gonorrhoeae (strain ATCC 700825 / FA 1090)
Neisseria gonorrhoeae (strain ATCC 700825 / FA 1090)
Thermus thermophilus (strain HB8 / ATCC 27634 / DSM 579)
Thermus thermophilus (strain HB8 / ATCC 27634 / DSM 579)
Thermus thermophilus (strain HB8 / ATCC 27634 / DSM 579)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
39000
-
isoform A, SDS-PAGE
45000
-
isoform B, SDS-PAGE
47000
-
isoform C, SDS-PAGE
84000
-
gel filtration
91000
gel filtration
105000
-
gel filtration
108600
isozyme GCYH-1B, apparent molecular weight estimated from gel filtration
120000
-
wild-type enzyme in presence of 0.3 M KCl, mutants C110G, E111K, H112D, and C181S, gel filtration
135000
-
gel filtration
170000
Serratia indica
-
gel filtration
200000
Serratia indica
-
gel filtration, D-I, D-II
210000
-
gel filtration
230000
-
gel filtration
250000
-
decameric wild-type enzyme, gel filtration
260000
-
GTP cyclohydrolase I
300000
-
larger than 300000, gel filtration
362000
-
gel filtration
400000
-
low molecular weight form, gel filtration
575000
-
gel filtration
600000
-
high molecular weight form, gel filtration
additional information
-
MW of mutant E152K is very high in gel filtration approximately corresponding to a didecameric form
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
-
x * 30000, SDS-PAGE
decamer
homodecamer
-
isozyme GCYH-1A
homodimer
homotetramer
octamer
Serratia indica
-
8 * 25000, SDS-PAGE
pentamer
-
GTP cyclohydrolase I feedback regulatory protein, i.e. GFRP
polymer
-
x * 39000, SDS-PAGE
tetramer
additional information
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
phosphoprotein
additional information
-
ubiquitination and proteasome-dependent degradation of GCH1. The CHIP-ubiquitin pathway modifies GCH1 in a lamb model of pulmonary hypertension secondary to increased pulmonary blood flow
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
selenomethionine-labeled isozyme GCYH-IB, sitting drop vapor diffusion method, using polyethylene glycol 6000 (10-16% (w/v)), LiCl (1-1.4 M), Tris (50 mM, pH 9.0), and Tris-HCl (50 mM, pH 7.0)
-
crystallization of purified recombinant enzyme mutants H112S, H113S, and C181S, free, mutant H112S in 0.1 M MES, pH 6.0, 0.2 M sodium acetate, 3 mM sodium azide, or complexed with substrate GTP, mutants H112S and C181S in 0.1 M MOPS, pH 7.0, 10% w/v PEG 6000, 0.1 M ammonium sulfate, or mutant H113S in 0.1 M Tris-HCl, pH 8.5, 0.2 M (NH4)H2PO4, 50% v/v 2-methyl-2,4-pentanediol, addition of GTP for the complex formation, hanging drop vapour diffusion method at room temperature, X-ray diffraction structure determination and analysis at about 2.1-3.2 A resolution, modeling
4.6 mg/ml purified recombinant DELTA42-hGTP-CH-I in 0.1 M potassium phosphate, pH 7.0, 0.02% NaN3, mixed with precipitation solution containing 6% PEG 6000, 150 mM KCl, 100 mM MOPS, pH 7.0, equilibration against precipitant solution, X-ray diffraction strcuture determination and analysis at 3.1 A resolution, modeling
-
5 mg/ml enzyme subunit GTP cyclohydrolase I feedback regulatory protein, i.e. GFRP, free and complexed with the human catalytic subunit of the enzyme, batch procedure, at 4C overnight, total reflection X-ray fluorescence spectrometry, structure determination and analysis at 2.6 A resolution, modeling
-
crystallization of purified recombinant enzyme and protein GFRP forming a BH2-induced inhibitory complex, involving dGTP, in 14% isopropanol, 0.2 M ammonium sulfate, 10% PEG 300, 10% ethylene glycol, 0.1 M MES-Na, pH 6.0, rapid freezing of crystals in liquid nitrogen, X-ray diffraction structure determination and analysis at 2.8 A resolution
-
purified recombinant enzyme complexed with recombinant wild-type and selenomethionine derivatized GFRP, the complex with the latter being used as CH3HGCl derivative, 5 mg/ml protein complex in 24% v/v 2-methyl-2,4-pentanediol, 75 mM Tris-HCl, pH 7.5, 50 mM KCl, 5 mM phenylalanine, X-ray diffraction structure determination and analysis at 2.7-2.8 A resolution, model building
-
purified enzyme complexed with 8-oxo-GTP or 8-oxo-dGTP, and as free enzyme, hanging drop vapour diffusion method, 20C, 0.002 ml of 13.7 mg/ml protein is mixed with 0.002 ml reservoir solution containing 0.1 M HEPES, pH 6.8, 2.0 M ammonium sulfate, 3.4% PEG 400, and 15% glycerol, X-ray diffraction structure determination and analysis at 2.0 and 1.8 A or at 2.2. A resolution, respectively
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
8.4
-
stability maximum
209552
additional information
-
-
209552
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50
-
88% of activity after 10 min
70 - 100
-
relative to a sample of the enzyme heated at 70C for 10 min, MptA lost 27% activity when it is incubated or 10 min at 80C, 60% activity at 90C, and 89% activity at 100C
80
-
human liver enzyme, half-life: 2 min
82
-
half-life: 7 min
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
bovine serum albumin stabilizes
-
ORGANIC SOLVENT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dithiothreitol
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-80C, 1 year, stable
-
-80C, 6 months, stable
4C, extreme instability of purified enzyme
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate precipitation, Ni-NTA column chromatography, and Sephacryl S-300HR gel filtration
-
expression in Escherichia coli
-
expression of wild-type and mutant proteins in Escherichia coli
-
GCH1 in complexes with cellular proteins from organs liver, heart, brain, and kidney, by immunoaffinity chromatography, isolation of mitochondria from liver, heart and kidney
-
Mono Q HR column chromatography
-
Ni-NTA resin column chromatography
-
Ni2+-Sepharose column chromatography, and gel filtration
O9701
recombinant DELTA42-hGTP-CH-I from Escherichia coli M15
-
recombinant enzyme from Escherichia coli strain BL21(DE3) by ion exchange chromatography
recombinant HAT-tagged enzyme from Escherichia coli, 6.8fold
recombinant His-tagged wild-type GTPCH maltose-binding protein fusion protein from Escherichia coli by amylose affinity chromatography and cleavage of the fusion tag by TEV protease, purification of His-tagged DELTA45-GTPCH mutant from HEK-293 cells by metal affinity chromatography
-
recombinant maltose binding protein fusion enzyme fromn Escherichia coli strain DH5alpha
-
recombinant wild-type and mutant enzymes
-
two forms
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cloned into the yeast expression vector pVT103-U and introduced into yeast strain 971/6a
enzyme expression in Escherichia coli strain BL21(DE3)
expressed in Escherichia coli B834(DE3) cells
-
expressed in Escherichia coli BL21 cells
-
expressed in Escherichia coli BL21(DE3)pLysS cells
-
expressed in Escherichia coli BL21(DE3)RIL cells
O9701
expressed in Escherichia coli strain BL21-Codon Plus (DE3)-PIL
-
expressed in Escherichia coli, baby hamster kidney cells, and NIH-293 cells
-
expressed in HEK-293 cells
-
expressed in Mus musculus
-
expressed in Mus musculus microvascular endothelial cells
-
expressed in PC-12 cells; transient co-expression of the enzyme with estrogen receptors in PC12 cells
-
expression in Escherichia coli
expression of DELTA42-hGTP-CH-I in Escherichia coli M15
-
expression of GTPCH under the control of a 935-bp fragment of the mouse myosin heavy chain gene promoter
-
expression of His-tagged wild-type GTPCH as maltose-binding protein fusion protein in Escherichia coli, expression of a His-tagged GTPCH truncation mutant, devoid of 45 N-terminal amino acids, DELTA45-GTPCH, in HEK-293 cells
-
expression of the enzyme in Escherichia coli strain DH5alpha as maltose binding protein fusion protein
-
expression of the enzyme in transgenic Lycopersicon esculentum plants using the Agrobacterium tumefaciens transfection system
gene gch and adjacent regions, DNA and amino acid sequence determination and analysis, uncommon start codon TTG, ribosomal binding site, overexpression in Escherichia coli strains JM109 and BL21(DE3) as HAT-tagged enzyme, and expression in Mycobacterium smegmatis from plasmid
overexpression of wild-type and mutant enzymes from plasmids in strain DH5alpha
-
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
Dube3a has a co-transcriptional activation function for GCH1; Dube3a has a co-transcriptional activation function for GCH1; Dube3a has a co-transcriptional activation function for GCH1. Punch protein isoform B levels change as a result of Dube3a overexpression or loss of function in the fly brain
exposure to the NO donor spermine NONOate (0-0.1 mM) leads to an dose-dependent increase in GCH1 protein. GCH1 promoter activity is enhanced by spermine NONOate in a CREB-dependent manner
-
tetracycline stimulation substantially increases GCH-1 activity about 20fold
-
the autoinhibitory peptide provides a molecular mechanism for physiological up-regulation of GTPCH activity
-
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
S37E
-
the enzymatic activity is significantly higher than of wild type GTPCH isoform C
C110G
-
site-directed mutagenesis, 0.22% activity compared to the wild-type enzyme, increased temperature optimum
C110S
highly reduced activity in both reaction steps using GTP or 2-amino-5-formylamino-6-ribofuranosylamino-4(3H)-pyrimidinone triphosphate as substrate, reduced zinc content compared to the wild-type enzyme
E111K
-
site-directed mutagenesis, 3.1% activity compared to the wild-type enzyme, increased temperature optimum
E152K
-
site-directed mutagenesis, 0.06% activity compared to the wild-type enzyme, increased pH optimum, decreased temperature optimum
H112D
-
site-directed mutagenesis, 0.23% activity compared to the wild-type enzyme, decreased pH optimum, increased temperature optimum
H112S
determination of crystal structure, highly reduced activity in both reaction steps using GTP or 2-amino-5-formylamino-6-ribofuranosylamino-4(3H)-pyrimidinone triphosphate as substrate, reduced zinc content compared to the wild-type enzyme
H113N
-
site-directed mutagenesis, 67% activity compared to the wild-type enzyme, decreased pH optimum, increased temperature optimum
H113S
determination of crystal structure, highly reduced activity in both reaction steps using GTP or 2-amino-5-formylamino-6-ribofuranosylamino-4(3H)-pyrimidinone triphosphate as substrate, reduced zinc content compared to the wild-type enzyme
H179N
-
mutant H179N is not able to perform the whole reaction step, but can catalyzes the reversible formation of reaction intermediate 2-amino-5-formylamino-6-ribosylamino-4(3H)-pyrimidinone
H179Q
-
site-directed mutagenesis, 0.8% activity compared to the wild-type enzyme, increased temperature optimum
K136E
-
site-directed mutagenesis, 0.25% activity compared to the wild-type enzyme, increased temperature optimum
L134Q
-
site-directed mutagenesis, 1.85% activity compared to the wild-type enzyme, increased temperature optimum
R139C
-
site-directed mutagenesis, 29.3% activity compared to the wild-type enzyme, decreased pH optimum
R185G
-
site-directed mutagenesis, 2.9% activity compared to the wild-type enzyme, decreased temperature optimum
R56L
-
site-directed mutagenesis, 14% activity compared to the wild-type enzyme, increased temperature optimum
S135C
-
site-directed mutagenesis, 0.7% activity compared to the wild-type enzyme, decreased pH and temperature optimum
V150E
-
site-directed mutagenesis, 3.2% activity compared to the wild-type enzyme, slightly decreased pH optimum, increased temperature optimum
C265T
-
mutation results in the loss of the 177 C-terminal amino acids
G155S
-
the mutation is associated with Dopa-responsive dystonia in Chines Han population
M211I
-
low activity at 0.1 mM GTP
R184H
-
very low activity at 0.1 mM GTP
R88W
-
very low activity at 0.1 mM GTP
S81A
-
site-directed mutagenesis, the mutant shows enhanced interaction with GFRP both in the presence of BH4 and GTP compared to the wild-type enzyme
S81D
-
site-directed mutagenesis, phospho-mimetic mutant that shows increased enzyme activity, reduced binding to GFRP, and resistance to inhibition by GFRP compared to wild-type GTPCH-1
H101N
-
shows wild type activity
H200N
-
shows reduced enzymatic activity but no reduction in the amount of bound iron
H293N
-
shows reduced enzymatic activity but no reduction in the amount of bound iron
H295N
-
shows reduced enzymatic activity but no reduction in the amount of bound iron
H101N
-
shows wild type activity
-
H200N
-
shows reduced enzymatic activity but no reduction in the amount of bound iron
-
H293N
-
shows reduced enzymatic activity but no reduction in the amount of bound iron
-
H295N
-
shows reduced enzymatic activity but no reduction in the amount of bound iron
-
H279S
O9701
inactive
N88Y
O9701
the mutant shows about wild type activity
R230K
O9701
the mutant shows about wild type activity
S130A
-
phospho-defective mutant with significantly decreased GCH-1 activity
S72A
-
phospho-defective mutant with significantly decreased GCH-1 activity
T103A
-
phospho-defective mutant with significantly decreased GCH-1 activity
T231A
-
mutant with 23% increased GCH-1 activity but reduced GCH-1 nuclear localization and nuclear GCH-1 activity
T85A
-
phospho-defective mutant with 67% decreased GCH-1 activity
T91A
-
phospho-defective mutant with significantly decreased GCH-1 activity
additional information
Renatured/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
denaturation of the purified recombinant enzyme by 4 M guanidine hydrochloride for 30 min, refolding of denatured enzyme by 100fold dilution in presence of GroE, a chaperone protein, effects of ZnSO4 and EGTA on refolding and activity
-
reconstitution of dissociated mutant enzyme subunits to chimeric dimers from 2 monomers A and B derived from 2 different mutants, overview
-
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
biotechnology
medicine
molecular biology
mutations in Punch can act as genetic enhancers of Dube3a over-expression phenotypes; mutations in Punch can act as genetic enhancers of Dube3a overexpression phenotypes
nutrition
construction of transgenic tomato plants expressing GCHI for engineering of the peteridine branch of folate synthesis in Lycopersicon esculentum by folate biofortification, overview
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