Information on EC 6.3.2.25 - Tubulin-tyrosine ligase

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The expected taxonomic range for this enzyme is: Eukaryota

EC NUMBER
COMMENTARY
6.3.2.25
-
RECOMMENDED NAME
GeneOntology No.
Tubulin-tyrosine ligase
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
ATP + detyrosinated alpha-tubulin + L-tyrosine = alpha-tubulin + ADP + phosphate
show the reaction diagram
-
-
-
-
ATP + detyrosinated alpha-tubulin + L-tyrosine = alpha-tubulin + ADP + phosphate
show the reaction diagram
random sequential mechanism
-
REACTION TYPE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
formation of peptide bond
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
alpha-Tubulin:L-tyrosine ligase (ADP-forming)
L-Tyrosine is linked via a peptide bond to the C-terminus of de-tyrosinated alpha-tubulin (des-Tyromega-alpha-tubulin). The enzyme is highly specific for alpha-tubulin and moderately specific for ATP and L-tyrosine. L-Phenylalanine and 3,4-dihydroxy-L-phenylalanine are transferred but with higher Km values.
SYNONYMS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
Synthetase, tubulin-tyrosine
-
-
-
-
TTL
-
-
-
-
TTL
Q9QXJ0
-
TTLase
-
-
-
-
tubulin tyrosine ligase
-
-
tubulin tyrosine ligase
-
-
tubulin tyrosine ligase
Q9QXJ0
-
tubulin tyrosine ligase
-
-
tubulin-tyrosine-ligase
-
-
Tubulin:tyrosine ligase
-
-
-
-
tubuline-tyrosine ligase
-
-
Tyrosyltubulin ligase
-
-
-
-
CAS REGISTRY NUMBER
COMMENTARY
60321-03-1
-
GENERAL INFORMATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
physiological function
-
plays a role in cell division, in particular in cell wall deposition
SUBSTRATE
PRODUCT                      
REACTION DIAGRAM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ATP + detyrosinated alpha-tubulin + 3-fluoro-Tyr
?
show the reaction diagram
-
tubulin covalently labeled with 3-fluoro-Tyr is competient to polimerize into microtubules
-
-
-
ATP + detyrosinated alpha-tubulin + 3-nitro-L-tyrosine
nitrotyrosinated alpha-tubulin + ADP + phosphate
show the reaction diagram
-
-
-
-
ir
ATP + detyrosinated alpha-tubulin + 3-nitro-L-tyrosine
nitrotyrosinated alpha-tubulin + ADP + phosphate
show the reaction diagram
-
35fold lower affinity for nitrotyrosine than for L-tyrosine, nitrotyrosine is incoorporated into the C-terminus
-
?
ATP + detyrosinated alpha-tubulin + iodotyrosine
?
show the reaction diagram
-
-
-
-
-
ATP + detyrosinated alpha-tubulin + L-DOPA
?
show the reaction diagram
-
-
-
-
-
ATP + detyrosinated alpha-tubulin + L-hydroxyphenylalanine
?
show the reaction diagram
-
-
-
-
-
ATP + detyrosinated alpha-tubulin + L-Phe
?
show the reaction diagram
-
-
-
-
-
ATP + detyrosinated alpha-tubulin + L-Phe
?
show the reaction diagram
-
-
-
-
-
ATP + detyrosinated alpha-tubulin + L-Tyr
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
-
-
-
ATP + detyrosinated alpha-tubulin + L-Tyr
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
-
-
-
-
ATP + detyrosinated alpha-tubulin + L-Tyr
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
r
-
-
-
ATP + detyrosinated alpha-tubulin + L-Tyr
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
tyrosinates Crithidia tubulin but not brain tubulin and is specific for the alpha chain
-
-
-
ATP + detyrosinated alpha-tubulin + L-Tyr
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
turkey erythrocyte tubulin
-
-
-
ATP + detyrosinated alpha-tubulin + L-Tyr
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
the enzyme plays a vital role in neuronal organization
-
-
?
ATP + detyrosinated alpha-tubulin + L-Tyr
alpha-tubulin + ADP + phosphate
show the reaction diagram
Q8NG68
the enzyme plays a vital role in neuronal organization
-
-
?
ATP + detyrosinated alpha-tubulin + L-Tyr
?
show the reaction diagram
-
activity increases from the embryonic stage to the fourth larval stage L4, decreases from L4 to adult and to aged nematodes with only qualitative alterations in substrate specificity
-
-
-
ATP + detyrosinated alpha-tubulin + L-Tyr
?
show the reaction diagram
-
posttranslational addition of Tyr to the carboxyterminal end of detyrosinated alpha-tubulin
-
-
-
ATP + detyrosinated alpha-tubulin + L-Tyr
?
show the reaction diagram
-
terminal Tyr is present in about 5% of flagellar alpha-chain from cells in stationary phase and 20% from cells from midlog phase. None is detected in tubulin from cytosol or the subpellicular corset. In contrast to mammalian cells, in which higher state of tyrosinylation characterizes recently assembled or unstable microtubules, terminal Tyr is present only in the most stable polymer, the flagellar double microtubules
-
-
-
ATP + detyrosinated alpha-tubulin + L-Tyr
?
show the reaction diagram
-
posttranslational modification of tubulin with potential significance for the regulation of microtubule assembly or function
-
-
-
ATP + detyrosinated alpha-tubulin + L-tyrosine
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
-
-
-
?
ATP + detyrosinated alpha-tubulin + L-tyrosine
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
-
-
?
ATP + detyrosinated alpha-tubulin + L-tyrosine
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
-
-
?
ATP + detyrosinated alpha-tubulin + L-tyrosine
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
-
-
?
ATP + detyrosinated alpha-tubulin + L-tyrosine
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
-
-
?
ATP + detyrosinated alpha-tubulin + L-tyrosine
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
-
-
?
ATP + detyrosinated alpha-tubulin + L-tyrosine
alpha-tubulin + ADP + phosphate
show the reaction diagram
Q9QXJ0
-
-
?
ATP + detyrosinated alpha-tubulin + L-tyrosine
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
tubulin tyrosination is a major factor affecting the recruitment of cytoskeleton-associated protein glycine-rich proteins at microtubule plus ends
-
-
?
ATP + detyrosinated alpha-tubulin + tyramine
?
show the reaction diagram
-
-
-
-
-
ATP + detyrosinated alpha-tubulin + tyramine
?
show the reaction diagram
-
very low activity
-
-
-
ATP + peptide of detyrosinated alpha tubulin + Tyr
peptide of alpha-tubulin + ADP + phosphate
show the reaction diagram
-
peptides ending like alpha-tubulin with the sequence Gly-Glu-Glu are optimally tyrosinated once a peptide length of 12 residues is reached. The carboxy-terminal tetradecapeptide of detyrosinated alpha-tubulin acts as substrate at 50fold lower efficiency than alphabeta-tubulin
-
-
-
additional information
?
-
-
the testis-specific apoptosis related gene TTL.6 (member of the tubulin-tyrosine ligase family) undergoes adaptive evolution in the lineage leading to humans
-
-
-
additional information
?
-
-
microtubule-associated protein 1B interaction with tubulin tyrosine ligase contributes to the control of microtubule tyrosination, interaction between MAP1B and TTL is not regulated by MAP1B phosphorylation, overview
-
-
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
ATP + detyrosinated alpha-tubulin + L-Tyr
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
the enzyme plays a vital role in neuronal organization
-
-
?
ATP + detyrosinated alpha-tubulin + L-Tyr
alpha-tubulin + ADP + phosphate
show the reaction diagram
Q8NG68
the enzyme plays a vital role in neuronal organization
-
-
?
ATP + detyrosinated alpha-tubulin + L-Tyr
?
show the reaction diagram
-
activity increases from the embryonic stage to the fourth larval stage L4, decreases from L4 to adult and to aged nematodes with only qualitative alterations in substrate specificity
-
-
-
ATP + detyrosinated alpha-tubulin + L-Tyr
?
show the reaction diagram
-
posttranslational addition of Tyr to the carboxyterminal end of detyrosinated alpha-tubulin
-
-
-
ATP + detyrosinated alpha-tubulin + L-Tyr
?
show the reaction diagram
-
terminal Tyr is present in about 5% of flagellar alpha-chain from cells in stationary phase and 20% from cells from midlog phase. None is detected in tubulin from cytosol or the subpellicular corset. In contrast to mammalian cells, in which higher state of tyrosinylation characterizes recently assembled or unstable microtubules, terminal Tyr is present only in the most stable polymer, the flagellar double microtubules
-
-
-
ATP + detyrosinated alpha-tubulin + L-Tyr
?
show the reaction diagram
-
posttranslational modification of tubulin with potential significance for the regulation of microtubule assembly or function
-
-
-
ATP + detyrosinated alpha-tubulin + L-tyrosine
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
-
-
-
?
ATP + detyrosinated alpha-tubulin + L-tyrosine
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
-
-
?
ATP + detyrosinated alpha-tubulin + L-tyrosine
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
-
-
?
ATP + detyrosinated alpha-tubulin + L-tyrosine
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
-
-
?
ATP + detyrosinated alpha-tubulin + L-tyrosine
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
-
-
?
ATP + detyrosinated alpha-tubulin + L-tyrosine
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
-
-
?
ATP + detyrosinated alpha-tubulin + L-tyrosine
alpha-tubulin + ADP + phosphate
show the reaction diagram
Q9QXJ0
-
-
?
ATP + detyrosinated alpha-tubulin + L-tyrosine
alpha-tubulin + ADP + phosphate
show the reaction diagram
-
tubulin tyrosination is a major factor affecting the recruitment of cytoskeleton-associated protein glycine-rich proteins at microtubule plus ends
-
-
?
additional information
?
-
-
the testis-specific apoptosis related gene TTL.6 (member of the tubulin-tyrosine ligase family) undergoes adaptive evolution in the lineage leading to humans
-
-
-
additional information
?
-
-
microtubule-associated protein 1B interaction with tubulin tyrosine ligase contributes to the control of microtubule tyrosination, interaction between MAP1B and TTL is not regulated by MAP1B phosphorylation, overview
-
-
-
COFACTOR
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
Ca2+
-
required for the N-formyl-Met-Leu-Phe-induced and the antibiotic A23187-induced stimulation
KCl
-
optimal concentration for the Tyr releasing activity: 10-30 mM
KCl
-
optimal concentration: 0.15-0.3 mM
KCl
-
required
KCl
-
-
Mg2+
-
required for activity
Mg2+
-
-
MgCl2
-
optimal concentration for the Tyr releasing activity: 3-20 mM
MgCl2
-
optimal concentration: 5-30 mM
MgCl2
-
required
MgCl2
-
-
INHIBITORS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
(1R,4aS,6R,8S)-6,8-dihydroxy-5-[(3E)-5-hydroxy-5-(3-hydroxy-5-oxo-2,5-dihydrofuran-2-yl)-3-methylpent-3-en-1-yl]-4a,6-dimethyldecahydronaphthalene-1-carboxylic acid
-
increases the non-tyrosinatable form of alpha-tubulin in human cancer cell cultures indicating to act as an inhibitor of TTL
3-nitro-L-tyrosine
-
50% inhibition of tyrosine incorporation at 50fold higher concentrations of nitrotyrosine
ADP
-
competitive with respect to ATP, non-competitive with respect to tubulin and L-Tyr
alpha-L-Glu-L-Tyr
-
-
diiodotyrosine
-
-
L-Ala-L-Tyr
-
-
L-hydroxyphenylalanine
-
-
L-Tyr-Gly
-
-
L-Tyr-L-Ala
-
-
L-Tyr-L-Glu
-
-
L-Tyr-L-Tyr
-
-
NEM
-
substrate protects
p-Chloromercuriphenyl sulfonic acid
-
-
p-Iodophenylalanine
-
-
phosphate
-
very weak
tyramine
-
-
Tyrosinated tubulin
-
very weak
-
monoiodotyrosine
-
-
additional information
-
no inhibition by thyronine and its iodinated derivatives
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
A23187
-
Ca2+ is required for the N-formyl-Met-Leu-Phe-induced and the A23187-induced stimulation
N-Formyl-Met-Leu-Phe
-
Ca2+ is required for the N-formyl-Met-Leu-Phe-induced and the A23187-induced stimulation
KM VALUE [mM]
KM VALUE [mM] Maximum
SUBSTRATE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
0.014
-
ATP
-
-
0.186
-
L-Dopa
-
-
0.16
-
L-hydroxyphenylalanine
-
-
0.432
-
L-Phe
-
-
2.9
-
L-Phe
-
-
0.017
-
L-Tyr
-
-
0.0254
-
L-Tyr
-
-
0.045
-
L-Tyr
-
-
0.0068
-
L-tyrosine
-
37C
0.002
-
tubulin
-
below 0.002 mM
-
8
-
tyramine
-
-
0.0019
-
untyrosinated tubulin
-
-
-
Ki VALUE [mM]
Ki VALUE [mM] Maximum
INHIBITOR
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
0.24
-
3-nitro-L-tyrosine
-
37C
SPECIFIC ACTIVITY [µmol/min/mg]
SPECIFIC ACTIVITY MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
0.011
-
-
-
0.039
-
-
-
0.324
-
-
-
0.737
-
-
-
additional information
-
-
-
pH OPTIMUM
pH MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
8
-
-
-
pI VALUE
pI VALUE MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
6.51
-
-
predicted from nucleotide sequence
LOCALIZATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
MOLECULAR WEIGHT MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
35000
-
-
gel filtration, MW of enzyme in crude extract, the MW of the purified enzyme is 150000. Addition of pure dimeric tubulin to the 35000 MW enzyme converts it back to the larger form which is apparently a stoichiometric 1:1 complex of tubulin and the 35000 MW enzyme
37000
-
-
gel filtration
40000
-
-
glycerol density gradient centrifugation. The MW of the ligase preincubated with excess phosphocellulose-tubulin is 150000
43000
-
-
glycerol gradient centrifugation
45000
-
-
loaded with binding partners, mass spectrometry
additional information
-
-
two-domain structure: the two domains interact tightly under physiological conditions. The 30000 MW domain carries the binding sites for beta-tubulin and ATP. The 14000 MW domain can possibly form an additional part of the catalytic site
SUBUNITS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
?
-
x * 43000, SDS-PAGE, immunoblot
monomer
-
1 * 46000, SDS-PAGE
monomer
-
1 * 40000, SDS-PAGE
monomer
-
x * 43425, calculation from nucleotide sequence
monomer
-
1 * 43425, deduced from nucleotide sequence
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
phosphoprotein
-
TTL sequence has at least 8 consensus phosphorylation sites to PKA, PKC, CKII, or tyrosine kinase, phosphorylation may be a potential mechanism for regulation of alpha-tubulin tyronisation
phosphoprotein
-
TTL contains a potential serine phosphorylation site for cAMP-dependent protein kinase
GENERAL STABILITY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
stabilized to dilution by addition of 0.1 mg/ml tubulin
-
photoinactivation in presence of 3-azido-L-Tyr and p-azido-L-Phe. PCMB protects by reversibly blocking essential thiol groups during illumination
-
STORAGE STABILITY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
-70C, stable for at least a year in 400 mM KCl with a protein concentration of about 1 mg/ml
-
-70C
-
-70C, 25 mM K+-morpholinoethanesulfonic acid, pH 6.8, 150 mM KCl, 12.5 MgCl2, 2.5 mM ATP, 1 mM dithiothreitol, 15% glycerol, at least 4 months, no loss of activity
-
-70C, in presence of 40% or 20% glycerol the enzyme can be stored for at least 6 months at a protein concentration of 0.02 mg/ml
-
-70C, protein concentration 0.02 mg/ml, 20-40% glycerol, stable for at least 1 year
-
Purification/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
recombinant GST-tagged enzyme from Escherichia coli strain BL21 by glutathione affinity chromatography
-
GST-TTL fusion protein, glutathione-sepharose affinity chromatography
-
Cloned/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
expression in HEK293T cells
Q8NG68
expression of GST-tagged TTL in Escherichia coli strain BL21, co-expression of TTL with MAP1B in COS-7 cells inducing an increase in tyrosinated microtubules
-
transient transfection of a TTL cDNA portion that encodes the beta-tubulin binding domain but lacks the catalytic domain into CHO-K1 cells
Q9QXJ0
cloning of cDNA
-
expression in Escherichia coli
-
expression of TTL in Escherichia coli and Sf9 insect cells
-
ENGINEERING
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
additional information
-
neurons lacking MAP1B, when exposed to drugs that reversibly depolymerize microtubules, do not fully recover tyrosinated microtubules upon drug removal
APPLICATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
analysis
-
utilization of ligase to determine the state of tyrosination of tubulin
synthesis
-
restoration of the excitability of the giant axon of Doryteuthis bleekeri by tubulin-tyrosine ligase and microtubule proteins