Information on EC 2.7.1.71 - shikimate kinase

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

EC NUMBER
COMMENTARY hide
2.7.1.71
-
RECOMMENDED NAME
GeneOntology No.
shikimate kinase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + shikimate = ADP + 3-phosphoshikimate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
phospho group transfer
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Biosynthesis of antibiotics
-
-
Biosynthesis of secondary metabolites
-
-
chorismate biosynthesis from 3-dehydroquinate
-
-
chorismate metabolism
-
-
Metabolic pathways
-
-
Phenylalanine, tyrosine and tryptophan biosynthesis
-
-
SYSTEMATIC NAME
IUBMB Comments
ATP:shikimate 3-phosphotransferase
-
CAS REGISTRY NUMBER
COMMENTARY hide
9031-51-0
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
-
Manually annotated by BRENDA team
gene aroK
-
-
Manually annotated by BRENDA team
strain K12
-
-
Manually annotated by BRENDA team
Hansenula henricii
-
-
-
Manually annotated by BRENDA team
strain 26695
Uniprot
Manually annotated by BRENDA team
strain SS1
-
-
Manually annotated by BRENDA team
enzyme aggregate contains 5 activities: EC 1.1.1.25, EC 2.7.1.71, EC 4.2.1.10, EC 4.2.3.4 and EC 2.5.1.19
-
-
Manually annotated by BRENDA team
-
SwissProt
Manually annotated by BRENDA team
Rhodosporidium sphaerocarpum
enzyme aggregate contains 5 activities: EC 1.1.1.25, EC 2.7.1.71, EC 4.2.1.10, EC 4.2.3.4 and EC 2.5.1.19
-
-
Manually annotated by BRENDA team
Rhodosporidium toruloides
enzyme aggregate contains 5 activities: EC 1.1.1.25, EC 2.7.1.71, EC 4.2.1.10, EC 4.2.3.4 and EC 2.5.1.19
-
-
Manually annotated by BRENDA team
enzyme aggregate contains 5 activities: EC 1.1.1.25, EC 2.7.1.71, EC 4.2.1.10, EC 4.2.3.4 and EC 2.5.1.19
-
-
Manually annotated by BRENDA team
Saccharomycopsis lipolytica
enzyme aggregate contains 5 activities: EC 1.1.1.25, EC 2.7.1.71, EC 4.2.1.10, EC 4.2.3.4 and EC 2.5.1.19
-
-
Manually annotated by BRENDA team
enzyme aggregate contains 5 activities: EC 1.1.1.25, EC 2.7.1.71, EC 4.2.1.10, EC 4.2.3.4 and EC 2.5.1.19
-
-
Manually annotated by BRENDA team
serotype 5b
-
-
Manually annotated by BRENDA team
serotype 5b
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
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'-deoxyATP + shikimate
2'-deoxyADP + shikimate 3-phosphate
show the reaction diagram
-
-
-
-
?
ATP + shikimate
ADP + 3-phosphoshikimate
show the reaction diagram
ATP + shikimate
ADP + shikimate 3-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
ATP + shikimate
ADP + 3-phosphoshikimate
show the reaction diagram
ATP + shikimate
ADP + shikimate 3-phosphate
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Cd2+
-
requirement for Mg2+ can partially be replaced by Mn2+, Ca2+, Co2+ and Cd2+
Cl-
-
increases the stability of the structure of the enzyme and influences the affinity of ADP for shikimate kinase
Ni2+
-
divalent cation required, Mg2+ and Mn2+ are most effective, Ca2+ activates to a lesser extent
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
(4R,7S,8aS)-4-[3-(morpholin-4-yl)-3-oxopropyl]-7-[[4-(trifluoromethoxy)benzyl]amino]hexahydropyrrolo[1,2-a]pyrazin-1(2H)-one
-
-
(4R,7S,8aS)-4-[3-oxo-3-(piperidin-1-yl)propyl]-7-[[4-(trifluoromethoxy)benzyl]amino]hexahydropyrrolo[1,2-a]pyrazin-1(2H)-one
-
-
(NH4)2SO4
-
5 mM, 29% inhibition
1-(4-chloro-2,5-dimethoxyphenyl)-1H-tetrazole-5-thiol
-
inhibitor identified by structure-based virtual screening, docking simulations
1-[(3-ethyl-2,6-dimethylquinolin-4-yl)sulfanyl]methanediamine
-
inhibitor identified by structure-based virtual screening, docking simulations
1-[(3S,5S)-5-[3-(1,3-benzodioxol-5-yl)-1,2,4-oxadiazol-5-yl]-1-methylpyrrolidin-3-yl]-3-propan-2-ylurea
-
-
2-(3-methyl-5-sulfanyl-4H-1,2,4-triazol-4-yl)-1-(1,2,3,4-tetrahydro-9H-carbazol-9-yl)ethanone
2-([[3-([(3R,4S)-4-[2-(4-methylpiperazin-1-yl)-2-oxoethyl]piperidin-3-yl]methyl)-1,2-oxazol-5-yl]methyl]carbamoyl)benzoic acid
-
-
2-[(6-methyl-4-oxo-3-phenyl-3,4,5,6,7,8-hexahydro[1]benzothieno[2,3-d]pyrimidin-2-yl)sulfanyl]acetamide
-
inhibitor identified by structure-based virtual screening, docking simulations
3-methoxy-4-[[2-([2-methoxy-4-[(4-oxo-2-thioxo-1,3-thiazolidin-5-ylidene)methyl]phenoxy]methyl)benzyl]oxy]benzaldehyde
-
noncompetitive inhibitor with respect to both shikimate and MgATP
3-phenyl-4-(5-sulfanyl-1H-tetrazol-1-yl)butanoic acid
-
inhibitor identified by structure-based virtual screening, docking simulations
3-[(8-methoxy-2-methyl-3-propylquinolin-4-yl)sulfanyl]propanoic acid
-
inhibitor identified by structure-based virtual screening, docking simulations
5-bromo-2-(5-[[1-(3,4-dichlorophenyl)-3,5-dioxo-4-pyrazolidinylidene]methyl]-2-furyl)benzoic acid
-
competitive inhibitor toward shikimate and noncompetitive inhibitor with respect to MgATP
5-[(6S)-5-[[5-(hydroxymethyl)furan-2-yl]methyl]-4,5,6,7-tetrahydro-3H-imidazo[4,5-c]pyridin-6-yl]-3-[4-(trifluoromethoxy)phenyl]-2H-1,2,4-oxadiazol-1-ium
-
-
5-[[(4-fluorobenzyl)sulfanyl]methyl]-4-methyl-4H-1,2,4-triazole-3-thiol
-
inhibitor identified by structure-based virtual screening, docking simulations
6-[3-(1,3-benzodioxol-5-yl)-1,2,4-oxadiazol-5-yl]-5-(3-phenylpropyl)-3a,4,5,6,7,7a-hexahydro-1H-imidazo[4,5-c]pyridine
-
-
AMP
-
4 mM, 27% inhibition
caffeic acid
-
1 mM, 44% inhibition
EDTA
-
-
ethyl 4-[([(6S)-6-[4-(propan-2-yl)furan-2-yl]-3,4,6,7-tetrahydro-5H-imidazo[4,5-c]pyridin-5-yl]carbonyl)amino]benzoate
-
-
N-(3-chloro-4-methylphenyl)-2-[(1-methyl-1H-tetrazol-5-yl)sulfanyl]acetamide
-
inhibitor identified by structure-based virtual screening, docking simulations
Na2SO4
-
5 mM, 27% inhibition
NaCl
-
250 mM, 50% inhibition
NSC162535
selective inhibitor, identification and binding analysis with enzyme mutant E144A by virtual docking analysis, isothermal titration calorimetry, and crystals structure analysis revealing an induced-fit mechanism, inactivation mechanism, detailed overview. Binding kinetics of wild-type and mutant enzymes
p-coumaric acid
-
1 mM, 14% inhibition
shikimate 3-phosphate
-
-
[(4-oxo-3-phenyl-3,4-dihydroquinazolin-2-yl)sulfanyl]acetic acid
-
inhibitor identified by structure-based virtual screening, docking simulations
[[1,3-dimethyl-9-(2-methylprop-2-en-1-yl)-2,6-dioxo-2,3,6,9-tetrahydro-1H-purin-8-yl]sulfanyl]acetic acid
-
inhibitor identified by structure-based virtual screening, docking simulations
additional information
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.048 - 0.67
ATP
0.039 - 20
shikimate
additional information
additional information
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
23 - 40
ATP
35 - 40
shikimate
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.00948
3-methoxy-4-[[2-([2-methoxy-4-[(4-oxo-2-thioxo-1,3-thiazolidin-5-ylidene)methyl]phenoxy]methyl)benzyl]oxy]benzaldehyde
-
in 100 mM TrisHCl (pH 8.0), 50 mM KCl, 5 mM MgCl2, 2 mM ATP, 2 mM phosphoenolpyruvate, 0.7 mM NADH, 3 U/ml proteinase K, 2.5 U/ml lactate dehydrogenase, and 2 mM shikimate, at 25C; pH 8.0, 25C
0.00219
5-bromo-2-(5-[[1-(3,4-dichlorophenyl)-3,5-dioxo-4-pyrazolidinylidene]methyl]-2-furyl)benzoic acid
-
in 100 mM TrisHCl (pH 8.0), 50 mM KCl, 5 mM MgCl2, 2 mM ATP, 2 mM phosphoenolpyruvate, 0.7 mM NADH, 3 U/ml proteinase K, 2.5 U/ml lactate dehydrogenase, and 2 mM shikimate, at 25C; pH 8.0, 25C
0.000005
asxe1
-
pH and temperature not specified in the publication
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0055
3-methoxy-4-[[2-([2-methoxy-4-[(4-oxo-2-thioxo-1,3-thiazolidin-5-ylidene)methyl]phenoxy]methyl)benzyl]oxy]benzaldehyde
Helicobacter pylori
-
in 100 mM TrisHCl (pH 8.0), 50 mM KCl, 5 mM MgCl2, 2 mM ATP, 2 mM phosphoenolpyruvate, 0.7 mM NADH, 3 U/ml proteinase K, 2.5 U/ml lactate dehydrogenase, and 2 mM shikimate, at 25C
0.0064
5-bromo-2-(5-[[1-(3,4-dichlorophenyl)-3,5-dioxo-4-pyrazolidinylidene]methyl]-2-furyl)benzoic acid
Helicobacter pylori
-
in 100 mM TrisHCl (pH 8.0), 50 mM KCl, 5 mM MgCl2, 2 mM ATP, 2 mM phosphoenolpyruvate, 0.7 mM NADH, 3 U/ml proteinase K, 2.5 U/ml lactate dehydrogenase, and 2 mM shikimate, at 25C
0.0049
NSC162535
Helicobacter pylori
P56073
pH 7.5, 25C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.05718
-
-
101.9
-
pH 6.8, 37C
410
-
isoenzyme SK1
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
8.6 - 9
-
-
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 9.5
-
pH 6.0: about 50% of maximal activity, pH 9.5: optimum
6.2 - 9
-
pH 6.2: about 20% of maximal activity in Tris propane buffer, pH 7.8: about 30% of maximal activity, pH 9.0: optimum
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30 - 37
-
assay at
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.61
-
sequence calculation
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
OsSK1 is induced by treatment with the elicitor N-acetylchitoheptaose; OsSK2 is induced by treatment with the elicitor N-acetylchitoheptaose
Manually annotated by BRENDA team
expression of OsSK1 is upregulated specifically during the heading stage of panicle development; expression of OsSK3 is upregulated specifically during the heading stage of panicle development
Manually annotated by BRENDA team
-
isoform SK2 is predominantly expressed early in embryogenesis and vegetative tissues throughout development, isoform SK1 is expressed near or below background levels in vegetative tissues and is only expressed at higher levels in mature embryos and senescing leaves
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Aquifex aeolicus (strain VF5)
Bacteroides thetaiotaomicron (strain ATCC 29148 / DSM 2079 / NCTC 10582 / E50 / VPI-5482)
Campylobacter jejuni subsp. jejuni serotype O:2 (strain ATCC 700819 / NCTC 11168)
Coxiella burnetii (strain RSA 493 / Nine Mile phase I)
Escherichia coli (strain K12)
Helicobacter pylori (strain ATCC 700392 / 26695)
Helicobacter pylori (strain ATCC 700392 / 26695)
Helicobacter pylori (strain ATCC 700392 / 26695)
Helicobacter pylori (strain ATCC 700392 / 26695)
Helicobacter pylori (strain ATCC 700392 / 26695)
Helicobacter pylori (strain ATCC 700392 / 26695)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
18580
-
calculated from amino acid sequence
19000
-
isoenzyme SK1, gel filtration
21400
-
isoenzyme SK2, gel filtration
28000
-
gel filtration
additional information
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
detailed structure-activity relationship analysis, overview
monomer
additional information
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
proteolytic modification
protein import into intact chloroplasts isolated from Pisum sativum seedlings reveals that the full-length form is translocated into chloroplasts and processed. NH2 -terminal sequence functions as chloroplast transit peptide; protein import into intact chloroplasts isolated from Pisum sativum seedlings reveals that the full-length form is translocated into chloroplasts and processed. NH2 -terminal sequence functions as chloroplast transit peptide; protein import into intact chloroplasts isolated from Pisum sativum seedlings reveals that the full-length form is translocated into chloroplasts and processed. NH2 -terminal sequence functions as chloroplast transit peptide
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
heat-stable isozyme AtSK2, X-ray diffraction structure determinaion and analysis at 2.35 A resolution
structural model based on structure of Mycobacterium tuberculosis enzyme. Molecular modeling and molecular dynamics simulation. Substrate shikimate binds to a pocket formed by the conserved residues Asp33, Arg57, Gly78 and Gly79, and Arg135
-
mutant enzyme K15M, sitting-drop vapor diffusion
-
vapor-diffusion method using NaCl as precipitant
-
hanging-drop vapor diffusion method. 1.8 A crystal structure of shikimate kinase. The crystal structure shows a three-layer alpha/beta fold consisting of a central sheet of five parallel beta-strands flanked by seven alpha-helices. An HpSK-shikimate-PO4 complex is also determined and refined to 2.3 A, revealing induced-fit movement from an open to a closed form on substrate binding; hanging drop vapour diffusion method using containing 0.2 M lithium sulfate, 30% (wt/vol) PEG 8000, and 0.1 M sodium acetate buffer (pH 6.5), at 20C
molecular modeling and docking of inhibitors. The active site is rather roomy and deep, forming an L-shape channel on the surface of the protein, and compound 3-methoxy-4-[[2-([2-methoxy-4-[(4-oxo-2-thioxo-1,3-thiazolidin-5-ylidene)methyl]phenoxy]methyl)benzyl]oxy]benzaldehyde prefers the corner area of L-shape channel, while compound 5-bromo-2-(5-[[1-(3,4-dichlorophenyl)-3,5-dioxo-4-pyrazolidinylidene]methyl]-2-furyl)benzoic acid binds the short arm of the channel in the binding interactions
-
wild-type dimeric enzyme, wild-type enzyme in complex with products ADP and shikimate 3-phosphate, enzyme mutant R57A, and enzyme mutant E114A in complex with selective inhibitor NSC162535, hanging drop vapour ddiffusion metho, 50 mg/ml protein in 40 mM Tris-HCl, pH 7.0, containing 100 mM NaCl mixed with an equal volume of reservoir solution and equilibrated against 0.06 ml of reservoir solution, containing 0.2 M Li2SO4, 30% w/v PEG 8000, and 0.1 M sodium acetate, pH 6.5 for the apo-enzyme, or containing 0.1 M HEPES sodium salt, pH 7.5, 0.1 M sodium acetate, 18% w/v PEG 8000, 2% w/v 2-propanol, and 5 mM shikimate and 5 mM MgATP for the product complex enzyme, or containing 0.1 M HEPES sodium salt, pH 8.0, 8% w/v 2-propanol and 18% w/v PEG 4000 for enzyme mutant R57A, or containing 0.1 M HEPES sodium salt, pH 6.7, and 1.2 M potassium sodium tartrate tetrahydrate for the enzyme mutant E114A with inhibitor, X-ray diffraction structure determination and analysis at 1.8 A, 2.3 A, 2.4 A, and 2.53 A resolution, respectively, molecular replacement
crystal structure of the enzyme complexed with MgADP and shikimic acid, determined at 2.3 A resolution, hanging-drop vapor-diffusion method
-
crystal structure of the enzyme in complex with MgADP- determined at 1.8 A resolution
-
hanging drop vapour diffusion method
-
hanging drop vapour diffusion method, enzyme in complex with ADP and shikimate in the absence of Mg2+ using 0.1 M Tris-HCl buffer pH 8.0, 17% PEG 1500 and 0.5-0.7 M LiCl, or enzyme in complex with ADP and Mg2+ in the absence of shikimate using 0.1 M Tris-HCl buffer pH 8.0, 20% PEG 3350 and 0.1 M MgCl2*6H2O; in complex with ADP-shikimate and with MgADP-, at 1.93 A and 2.8 A resolution, respectively. Presence of Mg2+ influences the conformation of the shikimate hydroxyl groups and the position of the side chains of some of the residues of the activesite. Presence of Cl- seems to influence the affinity of ADP and its position in the active site and the opening length of the LID domain. Shikimate binding causes a closing of the LID domain and also seems to influence the crystallographic packing
-
hanging-drop vapour-diffusion method. Crystal structure of shikimate kinase complexed with MgADP and shikimate determined at 2.3 A resolution
-
sitting drop vapour diffusion method using 0.2 M ammonium sulfate and 30% (w/v) poly(ethylene glycol) monomethyl ether 5000 in 0.1 M MES (pH 6.5) buffer
-
X-ray crystal structure of shikimate kinase with bound shikimate and adenosine diphosphate determined to a resolution of 2.15 A, sitting drop vapor diffusion method
-
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
37
isozyme AtSK2 is highly unstable and becomes inactivated at 37C, whereas the heat-induced isoform, AtSK1, is thermostable and fully active at 37C
40
-
midpoint of protein unfolding transition is 39.7C for the wild-type enzyme, 39C for the mutant enzyme C13S and 43.0C for the wild-type enzyme K15M in absence of ligands
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
differential scanning calorimetry experiments for evaluaton of the stability and unfolding of each of the enzyme mutants, overview
the enzyme is fully unfolded in 4 M urea
-
unfolding of the enzyme by guanidinium chloride, in the absence of ligands there is a loss of structure over the range of 1-3 M guanidinium chloride
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-18C, 30% v/v glycerol, stable for several months with gradual loss of activity
-
-20C, 50% glycerol, aggregate containing 5 activities: EC 1.1.1.25, EC 2.7.1.71, EC 4.2.1.10, 5-dehydroquinate synthase and 3-enoyl-pyruvylshikimate 5-phosphate synthase is stable for at least 1 month
-
-20C, in presence of 15% glycerol, stable for up to 4 weeks without loss of activity
-
-20C, protein concentration 1 mg/ml in 0.05% M Tris-HCl buffer, pH 7.5, 10 mM MgCl2, 0.1 M NaCl, 1 mM DTT, 10% loss of activity after 1 week, 90% loss of activity after 2 months
-
4C, 50% glycerol, aggregate containing 5 activities: EC 1.1.1.25, EC 2.7.1.71, EC 4.2.1.10, 5-dehydroquinate synthase and 3-enoyl-pyruvylshikimate 5-phosphate synthase is stable for at least 19 days
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
enzyme aggregate contains 5 activities: EC 1.1.1.25, EC 2.7.1.71, EC 4.2.1.10, EC 4.2.3.4 and EC 2.5.1.19
-
immobilized-nickel ion affinity chromatography and Sephacryl S-100 gel filtration
-
immobilized-nickel ion chromatography and Superdex-75 gel filtration
isoenzyme SK1 and SK2
-
partial
-
recombinant N-termminally His6-tagged wild-type and mutant enzymes from Escherichia coli strain BL21 (DE3) by immobilized metal affinity chromatography and dialysis
-
shikimate kinase II
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
aroK-endoded shikimate kinase, cloned and overexpressed in soluble form in Escherichia coli
-
cloning of aroK encoding shikimate kinase I. AroK protein, i.e. shikimate kinase I and AroL protein, i.e. shikimate kinase II are of comparable length and the homology between them extends the entire length of the two enzymes
-
expressed in Escherichia coli BL21(DE3) cells
-
expressed in Escherichia coli JM109 cells
expressed in Escherichia coli strain BL21(DE3)
-
expression of N-termminally His6-tagged wild-type and mutant enzymes in Escherichia coli strain BL21 (DE3)
-
gene expression patterns of isoforms, AtSK1 and AtSK2 showing a signature of regulatory subfunctionalization
generation of knockout and antisense strain from Escherichia coli parent strain DH5alpha
-
protein is expressed in Escherichia coli as N-terminal fusion with His6 tag and a TEV-protease cleavage site
-
wild-type and mutant enzyme expressed in Escherichia coli
-
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
expression of plant shikimate kinase is induced under specific conditions of environmental stress and developmental requirements in an isoform-specific manner
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
K138N
AtSK1 mutation relative to AtSK2 predicted to confer substantial stabilizing effect
K87R
AtSK1 mutation relative to AtSK2 predicted to confer substantial stabilizing effect
L233F
AtSK1 mutation relative to AtSK2 predicted to confer substantial stabilizing effect
N234K
AtSK1 mutation relative to AtSK2 predicted to confer substantial stabilizing effect
R120K
AtSK1 mutation relative to AtSK2 predicted to confer substantial stabilizing effect
S152N
AtSK1 mutation relative to AtSK2 predicted to confer substantial stabilizing effect
S251N
AtSK1 mutation relative to AtSK2 predicted to confer substantial stabilizing effect
T230S
AtSK1 mutation relative to AtSK2 predicted to confer substantial stabilizing effect
C13S
-
enzymatically active mutant, turnover-number is 65% of that of the wild-type enzyme
C162S
-
turnover-number is 1.14fold higher than that of the wild-type enzyme
D34N
-
inactive mutant enzyme
D33A
site-directed mutagensis, inactive mutant
D33E
site-directed mutagensis, inactive mutant
E114A
site-directed mutagensis, the mutant shows 82% of wlld-type activity
F48A
site-directed mutagensis, inactive mutant
F48Y
site-directed mutagensis, the mutant shows 40% of wlld-type activity
M10A
site-directed mutagensis, the mutant shows 38% of wlld-type activity
R116A
site-directed mutagensis, inactive mutant
R116K
site-directed mutagensis, inactive mutant
R132A
site-directed mutagensis, the mutant shows 5% of wlld-type activity
R132K
site-directed mutagensis, inactive mutant
R57A
site-directed mutagensis, the mutant shows 2% of wlld-type activity
R57K
site-directed mutagensis, the mutant shows 2% of wlld-type activity
K15I
-
site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme
K15R
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site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme
R110A
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site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme
R117
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site-directed mutagenesis, inactive mutant
T17I
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site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme
T17R
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site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme
additional information
Renatured/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
the enzyme is fully unfolded in 4 M urea. Approximately 95% of the enzyme activity can be recovered on dilution of the urea from 4 to 0.36 M. Refolding occurs in at least four kinetic phases, the slowest of which corresponds with the regain of shikimate binding and enzyme activity
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when the enzyme is unfolded by incubation in 4 M urea, addition of NaCl or Na2SO4 leads to relatively slow refolding of the enzyme. The refolded enzyme can bind shikimate, though more weakly than the native enzyme. The refolded enzyme does not appear to be capable of binding nucleotides, nor does it possess detectable catalytic activity. The refolding process brought about by addition of salt in the presence of 4 M urea is not associated with any change in the fluorescence of the probe 8-anilino-1-naphthalenesulfonic acid
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
drug development
synthesis
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