Information on EC 2.7.11.7 - myosin-heavy-chain kinase

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The expected taxonomic range for this enzyme is: Eukaryota

EC NUMBER
COMMENTARY hide
2.7.11.7
-
RECOMMENDED NAME
GeneOntology No.
myosin-heavy-chain kinase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + [myosin heavy-chain] = ADP + [myosin heavy-chain] phosphate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
phospho group transfer
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
ATP:[myosin heavy-chain] O-phosphotransferase
The enzyme from Dictyostelium sp. (slime moulds) brings about phosphorylation of the heavy chains of Dictyostelium myosin, inhibiting the actin-activated ATPase activity of the myosin. One threonine residue in each heavy chain acts as acceptor. While the enzyme from some species is activated by actin, in other cases Ca2+/calmodulin are required for activity.
CAS REGISTRY NUMBER
COMMENTARY hide
64763-54-8
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
-
Manually annotated by BRENDA team
strain Ax-3
-
-
Manually annotated by BRENDA team
strain AX2
SwissProt
Manually annotated by BRENDA team
strain AX3
SwissProt
Manually annotated by BRENDA team
strain JH10
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
malfunction
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
ATP + AKRVSMMR
ADP + AKRVS(-phosphate)MMR
show the reaction diagram
peptide derived from myosin I heavy chain kinase amino acid residues 4-11, exchange at position 4: Ser to Ala
phophorylation site is Ser8
?
ATP + caldesmon
ADP + caldesmon phosphate
show the reaction diagram
O76739, P42527, P90648
catalytic domain
-
-
?
ATP + casein
ADP + casein phosphate
show the reaction diagram
ATP + chicken gizzard myosin light chain
ADP + chicken gizzard myosin light chain phosphate
show the reaction diagram
ATP + GRGRSSVYS
ADP + GRGRSS(-phosphate)VYS
show the reaction diagram
ATP + GRSARVSTYA
ADP + GRSARVS(-phosphate)TYA
show the reaction diagram
peptide derived from Dictyostelium myosin ID
-
?
ATP + histone 2A
ADP + histone 2A phosphate
show the reaction diagram
ATP + MH1 peptide
ADP + MH1 peeptide phosphate
show the reaction diagram
-
-
-
?
ATP + myelin basic protein
ADP + myelin basic protein phosphate
show the reaction diagram
O76739, P42527, P90648
catalytic domain
-
-
?
ATP + myosin heavy chain
ADP + myosin heavy chain phosphate
show the reaction diagram
ATP + myosin heavy chain kinase
ADP + myosin heavy chain kinase phosphate
show the reaction diagram
ATP + myosin heavy-chain
ADP + myosin heavy chain phosphate
show the reaction diagram
ATP + myosin I heavy chain
ADP + myosin I heavy chain phosphate
show the reaction diagram
ATP + myosin II heavy chain
ADP + myosin II heavy chain phosphate
show the reaction diagram
ATP + myosin regulatory light chain
ADP + myosin regulatory light chain phosphate
show the reaction diagram
O76739, P42527, P90648
catalytic domain
-
-
?
ATP + peptide LMM58 of heavy chain
?
show the reaction diagram
-
4 mol phosphate per mol, Thr-residues are phosphorylated
-
-
?
ATP + phosvitin
ADP + phosvitin phosphate
show the reaction diagram
ATP + RKKFGESEKTKTKEFL
?
show the reaction diagram
ATP + RKKFGESEKTKTKEFL-amide
?
show the reaction diagram
ATP + smooth muscle myosin light chain
ADP + smooth muscle myosin light chain phosphate
show the reaction diagram
-
-
-
-
?
ATP + synthetic peptides
?
show the reaction diagram
ATP + troponin T
ADP + troponin T phosphate
show the reaction diagram
-
-
-
-
?
ATP + YAYDTRYRR
?
show the reaction diagram
ATP + [myosin heavy-chain]
ADP + [myosin heavy-chain] phosphate
show the reaction diagram
ATP + [myosin-heavy-chain]
ADP + [myosin-heavy-chain]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 + myosin heavy chain
ADP + myosin heavy chain phosphate
show the reaction diagram
ATP + myosin heavy chain kinase
ADP + myosin heavy chain kinase phosphate
show the reaction diagram
ATP + myosin heavy-chain
ADP + myosin heavy chain phosphate
show the reaction diagram
ATP + myosin I heavy chain
ADP + myosin I heavy chain phosphate
show the reaction diagram
ATP + myosin II heavy chain
ADP + myosin II heavy chain phosphate
show the reaction diagram
ATP + [myosin heavy-chain]
ADP + [myosin heavy-chain] phosphate
show the reaction diagram
ATP + [myosin-heavy-chain]
ADP + [myosin-heavy-chain]phosphate
show the reaction diagram
P42527
-
-
-
?
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Ca2+/calmodulin
heparin
-
-
Myosin I
-
inhibits autophosphorylation
-
NaCl
-
above 0.1 M
Positively charged polypeptides
-
strong inhibition, e.g. poly-(D-Lys), poly-(L-Lys), poly-(L-Arg) of different molecular weights
-
additional information
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
acidic phospholipids
DNA
-
autophosphorylation activity is increased 5-10fold in a Ca2+-independent manner
F-actin
G-actin
slight activation of MHCK A, activation is abolished by latrunculin A, which depolymerizes the actin filament
-
guanosine 5'-3-O-(thio)triphosphate-Rac1
10fold activation of autophosphorylation and kinase activity; functions cooperatively with acidic phospholipids to associate the enzyme with membranes; i.e. GTPgammaS-Rac1
-
heparin
-
autophosphorylation activity is increased 5-10fold in a Ca2+-independent manner
phosphatidylinositol
phosphatidylinositol 4,5-bisphosphate
10fold activation of autophosphorylation and kinase activity
phosphatidylserine
Phospholipid vesicles
additional information
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.098 - 0.116
AKRVSMMR
0.043 - 0.3
ATP
0.05 - 0.2
GRGRSSVYS
0.016 - 0.019
GRSARVSTYA
0.015
myosin heavy chain
-
pH 7.5, 22C
-
0.0003 - 0.0023
myosin IA
0.0002 - 0.0055
myosin IB
0.015
protein LMM58 heavy chain
-
pH 7.5, 22C
-
0.22
RKKFGESEKTKTKEFL
-
recombinant MHCK B, pH 7.0, 22C
-
0.1 - 0.28
RKKFGESEKTKTKEFL-amide
0.55
YAYDTRYRR
O76739, P42527, P90648
pH 7.0, 25C
-
additional information
additional information
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
5 - 5.4
AKRVSMMR
71 - 105
GRGRSSVYS
0.3 - 4.7
GRSARVSTYA
0.01 - 0.79
RKKFGESEKTKTKEFL-amide
14
YAYDTRYRR
Dictyostelium discoideum
O76739, P42527, P90648
pH 7.0, 25C
-
additional information
additional information
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
additional information
-
inhibition by positively charged polypeptides
-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.00032
-
partially purified enzyme
0.00063
-
purified enzyme
2.1
-
purified enzyme
3.03
-
purified enzyme, substrate histone 2A, 0.067 mM
4.6
-
purified enzyme, substrate myosin IB, 0.0024 mM
additional information
-
-
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 8.5
6 - 9
-
about half-maximal activity at pH 6.0, about 90% of maximal activity at pH 7.0 and pH 9.0
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
-
-
Manually annotated by BRENDA team
-
of the intestine, brush border cells
Manually annotated by BRENDA team
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
additional information
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
107000
-
gel filtration
160000
-
gel filtration
240000
-
gel filtration
490000
-
gel filtration, sucrose density gradient centrifugation
700000
-
above, gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
decamer
-
10 * 50000, SDS-PAGE, asymmetric complex with an axial ratio calculated for prolate ellipsoid of 6.1
monomer
additional information
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
phosphoprotein
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
MHCK A in complex with ATP and a peptide substrate, and MHCK A alpha kinase domain in individual complexes with AMP, ADP, and beta,gamma-methyleneadenosine 5'-triphosphate, X-ray diffraction structcure determination and analysis
-
purified recombinant wild-type and truncation mutant enzymes, hanging drop vapor diffusion method, mixing of 0.001 ml protein solution with 0.001 ml of reservoir solution containing 0.1 M Tris-HCl, pH 8.5, 0.2 M NaH2PO4, and 18% w/v PEG 8000, 4C, 10 days, X-ray diffraction structure determination and analysis at 1.9 A resolution
-
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
8
-
and above, unstable
640776
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25
-
1-2 h, stable
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
freeze-thawing inactivates
-
loses about 25% of activity for every freeze-thaw cycle
-
phosphorylated enzyme is somewhat less stable than unphosphorylated enzyme
-
very unstable if exposed to low salt, i.e. 50 mM KCl or less, in the absence of sucrose
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, in 25 mM HEPES buffer, pH 7.5, 1 mM DTT, 1 mM EDTA, 50 mM NaCl, 10% glycerol, or 20% sucrose, stable for 3 months
-
-20C, in 50% glycerol, 3 months stable and less than 20% loss of activity within 6 months
-
-20C, in 50% sucrose or glycerol, t1/2: 2 weeks
-
-20C, purified, stable up to 1 year
-
0C, at least 1 month
-
0C, further purified Ca2+/calmodulin-dependent isozyme, complete loss of activity within 1 day
-
0C, highly purified preparation of unphosphorylated enzyme, 25% loss of activity per day
-
0C, in 10 mM imidazole, pH 7, 100 mM KCl, 2 mM DTT, 60% sucrose, several weeks
-
0C, partially purified preparation of unphosphorylated enzyme, several days
-
0C, partly purified Ca2+/calmodulin-dependent isozyme, at least 2 weeks
-
4C, several months
-
indefinitely stable upon storage in liquid nitrogen, recovery of 60-70% activity after thawing
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
14000fold to near homogeneity
-
affinity chromatography using histone-resin
-
affinity chromatography using histone-resin; to near homogeneity
-
Ca2+/calmodulin-dependent heavy chain kinase, 84fold
-
enzyme copurifies with casein kinase II and Ca2+-independent myosin kinase
-
isolation of highly phosphorylated and unphosphorylated myosin II heavy chain kinase A
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MHCK B, recombinant from Dictyostelium discoideum cells as FLAG-tagged protein
-
recombinant GST-fusion catalytic domain of MHCK B, expressed in Escherichia coli; recombinant His-tagged catalytic domain of MHCK A, expressed in Escherichia coli; recombinant His-tagged catalytic domain of MHCK C, expressed in Escherichia coli
O76739, P42527, P90648
recombinant GST-tagged wild-type full-length and MHCKB truncation mutants by glutathione affinity chromatography
recombinant His-tagged catalytic domains, wild-type and mutants, from Sf9 insect cells
-
recombinant His-tagged full-length isozyme MHCK A and truncation mutant DELTAcoil-MHCK A from Dictyostelium discoideum to homogeneity by affinity chromatography, recombinant GST-fusion coiled-coil domain and catalytic domain from Escherichia coli to near homogeneity by affinity chromatography
recombinant His-tagged full-length isozyme MHCK A from Dictyostelium discoideum by affinity chromatography, recombinant GST-fusion truncation mutant DELTAcoil-MHCK A from Escherichia coli by affinity chromatography, the GST-tag is cleaved off by thrombin treatment
recombinant His-tagged or GFP-fusion proteins from Dictyostelium discoideum
recombinant His-tagged wild-type and mutant MHCK A alpha-kinase domains from Escherichia coli strain BL21(DE3) by nickel affinity chromatography
-
recombinant MHCK A GST-tagged catalytic domain by glutathione affinity chromatography; recombinant MHCK B GST-tagged catalytic domain by glutathione affinity chromatography
recombinant peptides of the enzymes N-terminus from Escherichia coli, His- or FLAG-tagged, and recombinant enzyme from Sf9 insect cells
-
separation of the Ca2+/calmodulin-dependent and -independent isozymes
-
solubilized by high-salt extraction, affinity chromatography, 4600fold, to homogeneity
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cells overexpressing GFP-GEF have both unphosphorylated and autophosphorylated MHCK A
-
cloning of wild-type and mutants of the catalytic domain of myosin I heavy chain kinase as His-tagged proteins and expression in Spodoptera frugiperda Sf9 cells via baculovirus infection system
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construction of His-tagged or FLAG-tagged N-terminal enzyme peptides by site directed mutagenesis for introduction of start and stop codons, expression in Escherichia coli BL21(DE3); subcloning and expression in Escherichia coli, expression in Spodoptera frugiperda Sf9 cells via baculovirus infection system
-
construction of His-tagged or GFP fusion proteins with the full length enzyme or enzyme fragments, expression in Dictyostelium discoideum cells
DNA and amino acid sequence determination and analysis
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DNA sequence determination and analysis
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DNA sequence determination and analysis, gene MHCK possesses all of the domains characteristic of members of the protein kinase C family
expression of FLAG-tagged T825A and T825E enzyme mutants in Dictyostelium discoideum cells, expression of His-tagged wild-type and mutant MHCK A alpha-kinase domains in Escherichia coli strain BL21(DE3)
-
expression of GFP-tagged isozyme MHCK A in a Dictyostelium discoideum strain lacking endogenous MHCK A, expression of a truncation mutant DELTAcoil-MHCK A comprising residues 1-498 in Escherichia coli as GST-fused protein
expression of MHCK A, comprising residues 527-887, i.e. A-CAT, as a GST-fusion protein, phylogenetic analysis; expression of MHCK B, comprising residues 13-459, i.e. B-CAT, as a GST-fusion protein, phylogenetic analysis
expression of the 3 isozymes MHCK A, MHCK B, and MHCK C as GFP-fusion proteins
-
expression of the catalytic domain of MHCK B as a GST-fusion protein in Escherichia coli BL21(DE3); expression of the His-tagged catalytic domain of MHCK A in Escherichia coli BL21(DE3); expression of the His-tagged catalytic domain of MHCK C in Escherichia coli BL21(DE3)
O76739, P42527, P90648
gene mhck A, construction of overexpressing cell line by transfection of an expression plasmid into the deficient mhck A- cell line
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MHCK B, overexpression as FLAG-tagged protein in AX2 cells or 3xALA cells of Dictyostelium discoideum
-
overexpression of FLAG-tagged full-length MHCK-B in Dictyostelium cells, in AX2, mhkB-null, and mhkA/B/C-null cell lines. Expression of GST-tagged wild-type full-length and MHCKB truncation mutants
overexpression of His-tagged full-length isozyme MHCK A and truncation mutant DELTAcoil-MHCK A comprising residues 1-498 in Dictyostelium discoideum, overexpression of isolated coiled-coil domain and of the catalytic domain in Escherichia coli as GST-fused proteins
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
S627A
-
catalytic domain mutant, mutation of potential phosphorylation site, no phosphorylation of the mutant, reduced activity, increased Km values for the substrates, but higher activity than the unphosphorylated wild-type enzyme
S627D
-
catalytic domain mutant, mutation of potential phosphorylation site, no phosphorylation of the mutant, reduced activity, increased Km values for the substrates, the acidic residue cannot substitute for phospho-Ser
S627E
-
catalytic domain mutant, mutation of potential phosphorylation site, no phosphorylation of the mutant, reduced activity, increased Km values for the substrates, the acidic residue cannot substitute for phospho-Ser
T631A
-
catalytic domain mutant, mutation of a conserved Thr residue, full phosphorylation of the mutant, reduced activity, increased Km values for the substrates
T631D
-
catalytic domain mutant, mutation of a conserved Thr residue, 95% phosphorylation of the mutant, highly reduced activity, increased Km values for the substrates
T631E
-
catalytic domain mutant, mutation of a conserved Thr residue, no phosphorylation of the mutant, highly reduced activity, highly increased Km values for the substrates
T632A
-
catalytic domain mutant, mutation of a nonconserved Thr residue, full phosphorylation of the mutant, increased activity, only slightly increased Km values for the substrates
T632D
-
catalytic domain mutant, mutation of a nonconserved Thr residue, 80% phosphorylation of the mutant, reduced activity, increased Km values for the substrates
T632E
-
catalytic domain mutant, mutation of a nonconserved Thr residue, full phosphorylation of the mutant, reduced activity, increased Km values for the substrates
C800A
catalytic domain mutation, no remaining activity when expressed as full length enzyme or as catalytic domain only in a deficient Dictyostelium discoideum cell line
D766A
-
site-directed mutagenesis of the central active site residue, inactive mutant
D766E
-
site-directed mutagenesis of the central active site residue, inactive mutant
D766S
-
site-directed mutagenesis of the central active site residue, inactive mutant
K645A
-
site-directed mutagenesis of an alpha-kinase domain residue, the mutant shows 98.5% reduced activity compared to the wild-type enzyme
Q822R/Q823R/T825S
-
site-directed mutagenesis
R592A
-
site-directed mutagenesis of an alpha-kinase domain residue, the mutant shows 92% reduced activity compared to the wild-type enzyme
T825A
-
site-directed mutagenesis
T825E
-
site-directed mutagenesis
T825S
-
site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme
additional information
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