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Information on EC 3.4.21.66 - Thermitase and Organism(s) Thermoactinomyces vulgaris and UniProt Accession P04072
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3.4.21.66 ThermitaseSpecify your search results
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- 3.4.21.66
- subtilisins
-
3.4.21.14
-
carlsberg
-
subtilisin-like
-
subtilases
-
subtilisin-type
- eglin-c
- biotechnology
- analysis
The taxonomic range for the selected organisms is: Thermoactinomyces vulgaris
The enzyme appears in selected viruses and cellular organisms
The enzyme appears in selected viruses and cellular organisms
Synonyms
thermitase, thermostable serine protease, proteolysin, plla-degrading enzyme, thermoactinomyces vulgaris serine proteinase, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
Proteinase, Thermoactinomyces vulgaris serine
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-
-
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Thermoactinomyces vulgaris serine proteinase
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-
-
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Thermophilic Streptomyces serine proteinase
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-
-
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
hydrolysis of peptide bond
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-
-
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CAS REGISTRY NUMBER
COMMENTARY
69772-87-8
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SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
Anionic protease + H2O
?
-
at pH 8 or 6 and 25Ā°C or 4Ā°C completely hydrolyzed
-
-
?
Benzyloxycarbonyl-L-Ala-L-Ala-L-Leu 4-nitroanilide + H2O
?
-
-
-
-
?
Cry11A endotoxin + H2O
?
-
substrate from Bacillus thuringiensis ssp. israelensis, enzyme hydrolyzes the I28-A29 bond. At increasing concentration of enzyme, additionally lysis of M71-E72 bond
-
-
?
Urea-denatured hemoglobin + H2O
Hydrolyzed urea-denatured hemoglobin
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-
-
-
?
additional information
?
-
-
at the active site serine and histidine, cysteine and methionine are essential for activity
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-
?
additional information
?
-
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substrate specificity with proteins, amino acid esters, fatty acid esters and amino acid 4-nitroanilides
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-
?
additional information
?
-
-
thermitase has 5 subsites at the S-site of its active center
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-
?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Ca2+
Ca2+
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the enzyme binds very strongly one Ca2+, that cannot be removed without denaturation by any known method. A Ca2+ from a second binding site can be removed completely using Ca2+ chelating reagents
Ca2+
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three calcium binding sites, two of these are tight binding sites, and removal of calcium causes unfolding and autolysis to occur. The third calcium is more weakly bound, its removal reduces the activity of the enzyme by 10%, the reduction being reversible
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
2,2,5,5-Tetramethylpyrrolin-1-oxyl-3-carbonyl-Ala2-Phe methyl ketone
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2,2,5,5-Tetramethylpyrrolin-1-oxyl-3-carbonyl-Ala3-Phe methyl ketone
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Benzyloxycarbonyl-(Ala)n-Phe methyl ketone
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reacts irreversibly in a 1:1 ratio with the enzyme
Benzyloxycarbonyl-Ala2-Phe chloromethyl ketone
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and the corresponding methyl ketone
Dansyl-(Ala)n-PheCH2Cl
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n: 1, 2 and 3, and homologous carboxybenzoyl-protected peptides, with a higher inhibitory effect
Peptide diazomethyl ketones
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reacts irreversibly in a 1:1 ratio with the enzyme
Peptide methyl ketones
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kinetic studies on the binding of N-acetylated peptide ketones as substrate analogous
additional information
-
stereospecificity of the binding of peptide ligands, diastereomeric tetrapeptides of the peptide ketone, benzoyl-(L-Ala)3-L-PheCH3, a competitive inhibitor
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additional information
-
structure of the active centre studied by use of spin-labeled peptide methyl ketones
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
-
cysteine residues essential for activity
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
7 - 9
-
small differences between the individual high-MW and low-MW substrates
additional information
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
-
the temperature optimum increases with incresing MW of substrate
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). PDB
SCOP
CATH
UNIPROT
ORGANISM
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
28369
-
1 * 28369, Thermoactinomyces vulgaris, calculation from primary structure
28370
-
Thermoactinomyces vulgaris, calculation from primary structure
additional information
additional information
-
the large CNBr fragment represents the N-terminal part of the molecule and contains 266 amino acids, MW 22932
additional information
-
characterization of 2 peptides obtained by CNBr cleavage, MW 25000 and 26000
additional information
-
amino acid sequence of the tryptic SH-peptide
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
monomer
monomer
-
1 * 28369, Thermoactinomyces vulgaris, calculation from primary structure
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
a new crystal form of native thermitase has been obtained using formate as the precipitating agent
-
thermitase-eglin-c complex (at 1.98 A resolution, and comparison of 2 crystal forms that differ in calcium content)
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three-dimensional structure comparison of native thermitase and thermitase-eglin-c complexes
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PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
computational procedure OptGraft for placing a novel binding pocket onto a protein structure so as its geometry is minimally perturbed. Transfer of a calcium-binding pocket from thermitase protein, PDB entry 1thm, into the first domain of rat CD2 protein, PDB entry 1hng results in new proteins that all exhibit high affinities for terbium and can selectively bind calcium over magnesium
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
-
autolysis and thereby inactivation at alkaline pH
29549
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
additional information
-
calcium ions contribute to the overall stability of the surface regions and improve the thermal stability of the enzyme
additional information
-
unusually tight binding of Ca2+ by thermitase emerges as the most likely single influence responsible for its increased thermostability
additional information
-
irreversible inhibition by chloromethyl ketones causes marked stabilization against thermal denaturation
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
Anionic protease, a component which frequently contaminates preparations of routinely isolated cationic protease, stabilizes thermitase
-
Autolysis and thereby inactivation at elevated temperature, at alkaline pH-values and in the absence of added substrate
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Irreversible inhibition by chloromethyl ketones causes marked stabilization against thermal denaturation, reversible inhibitors have no influence on stability
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The stability is significantly improved by 1 M acetate and chloride
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Unusually tight binding of Ca by thermitase emerges as the most likely single influence responsible for its increased thermostability
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PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
biotechnology
computational procedure OptGraft for placing a novel binding pocket onto a protein structure so as its geometry is minimally perturbed. Transfer of a calcium-binding pocket from thermitase protein, PDB entry 1thm, into the first domain of rat CD2 protein, PDB entry 1hng results in new proteins that all exhibit high affinities for terbium and can selectively bind calcium over magnesium
analysis
-
use of enzyme for generation of peptides for mass spectrometric characterization
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Betzel, C.; Teplyakov, A.V.; Harutyunyan, E.H.; Saenger, W.; Wilson, K.S.
Thermitase and proteinase K: a comparison of the refined three-dimensional structures of the native enzymes
Protein Eng.
3
161-172
1990
Thermoactinomyces vulgaris
Meloun, B.; Baudys, M.; Kostka, V.; Hausdorf, G.; Frömmel, C.; Höhne, W.E.
Complete primary structure of thermitase from Thermoactinomyces vulgaris and its structural features related to the subtilisin- type proteinases
FEBS Lett.
183
195-200
1985
Thermoactinomyces vulgaris
-
Mizusawa, K.; Yoshida, F.
Thermophilic Streptomyces alkaline proteinase. I. Isolation, crystallization, and physicochemical properties
J. Biol. Chem.
247
6978-6984
1972
Streptomyces rectus var. proteolyticus, Thermoactinomyces vulgaris
Teplyakov, A.V.; Kuranova, I.P.; Harutyunyan, E.H.; Vainshtein, B.K.; Frömmel, C.; Höhne, W.E.; Wilson, K.S.
Crystal structure of thermitase at 1.4 A resolution
J. Mol. Biol.
214
261-279
1990
Thermoactinomyces vulgaris
Teplyakov, A.V.; Kuranova, I.P.; Harutyunyan, E.H.; Frömmel, C.; Höhne, W.E.
Crystal structure of thermitase from Thermoactinomyces vulgaris at 2.2 A resolution
FEBS Lett.
244
208-212
1989
Thermoactinomyces vulgaris
Gros, P.; Kalk, K.H.; Hol, W.G.J.
Calcium binding to thermitase. Crystallographic studies of thermitase at 0, 5, and 100 mM calcium
J. Biol. Chem.
266
2953-2961
1991
Thermoactinomyces vulgaris
Gros, P.; Betzel, C.; Dauter, Z.; Wilson, K.S.; Hol, W.G.J.
Molecular dynamics refinement of a thermitase-eglin-c complex at 1.98 A resolution and comparison of two crystal forms that differ in calcium content
J. Mol. Biol.
210
347-367
1989
Thermoactinomyces vulgaris
Kelders, H.A.; Kalk, K.H.; Hol, W.G.J.
Crystallization of thermitase, a thermostable subtilisin, from a sodium formate solution by means of an automated procedure
J. Mol. Biol.
205
615-616
1989
Thermoactinomyces vulgaris
Brömme, D.; Kleine, R.
Use of microbial peptide inhibitors for characterization of the substrate specificity of thermitase, a thermostable serine protease from Thermoactinomyces vulgaris
Curr. Microbiol.
11
317-320
1984
Thermoactinomyces vulgaris
-
Dauter, Z.; Betzel, C.; Höhne, W.E.; Ingelman, M.; Wilson, K.S.
Crystal structure of a complex between thermitase from Thermoactinomyces vulgaris and the leech inhibitor eglin
FEBS Lett.
236
171-178
1988
Thermoactinomyces vulgaris
Briedigkeit, L.; Frömmel, C.
Calcium ion binding by thermitase
FEBS Lett.
253
83-87
1989
Thermoactinomyces vulgaris
-
Kleine, R.; Kettmann, U.
Separation and comparative characterization of the cationic protease and anionic protease from the culture medium of Thermoactinomyces vulgaris
Hoppe-Seyler's Z. Physiol. Chem.
363
843-853
1982
Thermoactinomyces vulgaris
Stepanov, V.M.; Chestukhina, G.G.; Rudenskaya, G.N.; Epremyan, A.S.; Osterman, A.L.; Khodova, O.M.; Belyanova, L.P.
A new subfamily of microbial serine proteinase? Structural similarities of Bacillus thuringiensis and Thermoactinomyces vulgaris extracellular serine proteinases
Biochem. Biophys. Res. Commun.
100
1680-1687
1981
Thermoactinomyces vulgaris
Stepanov, V.M.; Revina, L.P.; Abramov, S.T.; Strongin, A.Y.; Behnke, U.
Affinity chromatography of thermitase
J. Appl. Biochem.
2
342-345
1980
Thermoactinomyces vulgaris
-
Betzel, C.; Rachev, R.; Dolashka, P.; Genov, N.
Actinomycins as proteinase inhibitors
Biochim. Biophys. Acta
1161
47-51
1993
Thermoactinomyces vulgaris
Behnke, U.; Ruttloff, H.; Kleine, R.
Preparation and characterization of proteases from Thermoactinomyces vulgaris. V. Investigations on autolysis and thermostability of the purified protease
Z. Allg. Mikrobiol.
22
511-519
1982
Thermoactinomyces vulgaris
Froemmel, C.; Hausdorf, G.; Hoehne, W.E.; Behnke, U.; Ruttloff, H.
Characterization of a protease from Thermoactinomyces vulgaris (thermitase). 2. Single-step fine purification and protein-chemical characterization
Acta Biol. Med. Ger.
37
1193-1204
1978
Thermoactinomyces vulgaris
Gros, P.; Teplyakov, A.V.; Hol, W.G.J.
Effects of eglin-c binding to thermitase: three-dimensional structure comparison of native thermitase and thermitase eglin-c complexes
Proteins Struct. Funct. Genet.
12
63-74
1992
Thermoactinomyces vulgaris
Ermer, A.; Baumann, H.; Steude, G.; Peters, K.; Fittkau, S.; Dolashka, P.; Genov, N.
Peptide diazomethyl ketones are inhibitors of subtilisin-type serine proteases
J. Enzyme Inhib.
4
35-42
1990
Thermoactinomyces vulgaris
Froemmel, C.; Sander, C.
Thermitase, a thermostable subtilisin: comparison of predicted and experimental structures and the molecular cause of thermostability
Proteins Struct. Funct. Genet.
5
22-37
1989
Thermoactinomyces vulgaris
Schaedlich, H.; Flemming, C.; Gabert, A.; Taeufel, A.
Purification by affinity chromatography and characterization of two stable thermitase inhibitors from potatoes
Nahrung
32
91-107
1988
Thermoactinomyces vulgaris
-
Fittkau, S.; Pauli, D.; Bouaravong, P.; Damerau, W.
Thermitase: a thermostable serine protease. VIII. Kinetic and ESR investigations on the interactions of enzymes with spin labeled peptide methyl ketones
Biomed. Biochim. Acta
45
877-886
1986
Thermoactinomyces vulgaris
Broemme, D.; Fittkau, S.
A new substrate and two inhibitors applicable for thermitase, subtilisin BPN and alpha-chymotrypsin. Comparison of kinetic parameters with customary substrates and inhibitors
Biomed. Biochim. Acta
44
1089-1094
1985
Thermoactinomyces vulgaris
Meloun, B.; Baudys, M.; Pavlik, M.; Kostka, V.; Hausdorf, G.; Hoehne, W.E.
Thermitase from Thermoactinomyces vulgaris; amino acid sequence of the large N-terminal cyanogen bromide peptide
Collect. Czech. Chem. Commun.
50
885-896
1985
Thermoactinomyces vulgaris
-
Fittkau, S.; Brutscheck, M.
Thermitase: a thermostable serine protease. V. Stereospecificity of the binding of peptide ligands
Biomed. Biochim. Acta
43
901-907
1984
Thermoactinomyces vulgaris
Fittkau, S.; Smalla, K.; Pauli, D.
Thermitase: a thermostable serine protease. IV. Kinetic studies on the binding of N-acyl peptide ketones as substrate analog inhibitors
Biomed. Biochim. Acta
43
887-899
1984
Thermoactinomyces vulgaris
Peters, K.; Baetz, O.; Hoehne, W.E.; Fittkau, S.
Thermitase - a thermostable serine protease. VI. Kinetic and fluorescence studies on the interaction of the enzymes with dansylated peptide chloromethylketones
Biomed. Biochim. Acta
43
909-918
1984
Thermoactinomyces vulgaris
Schreier, E.; Fittkau, S.; Hoehne, W.E.
Influence of synthetic peptide inhibitors on the thermal stability of thermitase, a serine proteinase from Thermoactinomyces vulgaris
Int. J. Pept. Protein Res.
23
134-141
1984
Thermoactinomyces vulgaris
Baudys, M.; Kostka, V.; Hausdorf, G.; Fittkau, S.; Hoehne, W.E.
Amino acid sequence of the tryptic SH-peptide of thermitase, a thermostable serine proteinase from Thermoactinomyces vulgaris. Relation to the subtilisins
Int. J. Pept. Protein Res.
22
66-72
1983
Thermoactinomyces vulgaris
Kleine, R.
Properties of thermitase, a thermostable serine protease from Thermoactinomyces vulgaris
Acta Biol. Med. Ger.
41
89-102
1982
Thermoactinomyces vulgaris
Hausdorf, G.; Krueger, K.; Hoehne, W.E.
Thermitase, a thermostable serine protease from Thermoactinomyces vulgaris. Classification as a subtilisin-type protease
Int. J. Pept. Protein Res.
15
420-429
1980
Thermoactinomyces vulgaris
Behnke, U.; Kleine, R.; Ludewig, M.; Ruttloff, H.
Characterization of a protease from Thermoactinomyces vulgaris (thermitase). 3. Substrate specificity and properties of partially purified thermitase
Acta Biol. Med. Ger.
37
1205-1214
1978
Thermoactinomyces vulgaris
Behnke, U.; Schalinatus, E.; Ruttloff, H.; Hoehne, W.E.; Froemmel, C.
Characterization of a protease from Thermoactinomyces vulgaris (thermitase). 1. Purification of thermitase
Acta Biol. Med. Ger.
37
1185-1192
1978
Thermoactinomyces vulgaris
Revina, L.P.; Kostina, L.I.; Ganushkina, L.A.; Mikhailova, A.L.; Zalunin, I.A.; Chestukhina, G.G.
Reconstruction of Bacillus thuringiensis ssp. israelensis Cry11a endotoxin from fragments corresponding to its N- and C-moieties restores its original biological activity
Biochemistry (Moscow)
69
181-187
2004
Thermoactinomyces vulgaris
Schmelzer, C.E.; Getie, M.; Neubert, R.H.
Mass spectrometric characterization of human skin elastin peptides produced by proteolytic digestion with pepsin and thermitase
J. Chromatogr. A
1083
120-126
2005
Thermoactinomyces vulgaris
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