Information on EC 3.4.21.B56 - Pyrococcus horikoshii membrane protease PH1510

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The expected taxonomic range for this enzyme is: Pyrococcus horikoshii

EC NUMBER
COMMENTARY hide
3.4.21.B56
preliminary BRENDA-supplied EC number
RECOMMENDED NAME
GeneOntology No.
Pyrococcus horikoshii membrane protease PH1510
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
N-terminal region of open reading frame, PH1510 (residues 16–236, designated as 1510-N) is a serine protease
UniProt
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
physiological function
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the enzyme may be involved in the quality control of membrane proteins, e.g. stomatin, prohibitin, flotillin, and HflK/C domain proteins found in the lipid raft microdomains of various cellular membranes
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
alpha-casein + H2O
?
show the reaction diagram
alpha-casein is not cleaved as effectively as beta-casein. 1510-N probably cleaves the substrate specifically. The cleavage sites contain many hydrophobic and aromatic residues. The 1510-N protease possibly recognizes leucine at the cleavage site
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?
beta-casein + H2O
?
show the reaction diagram
alpha-casein is not cleaved as effectively as beta-casein. 1510-N probably cleaves the substrate specifically. The cleavage sites contain many hydrophobic and aromatic residues. The 1510-N protease possibly recognizes leucine at the cleavage site
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?
p-stomatin PH1511 + H2O
?
show the reaction diagram
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the N-terminal region of PH1510 (residues 16-236, i.e. 1510-N) is a serine protease with a catalytic Ser-Lys dyad (Ser97 and Lys138) and specifically cleaves the C-terminal hydrophobic region of the p-stomatin PH1511
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?
p-stomatin PH1511p + H2O
?
show the reaction diagram
the N-terminal region of PH1510p is a serine protease and specifically cleaves the C-terminal hydrophobic region of the p-stomatin PH1511p
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?
stomatin-homolog PH1511 + H2O
?
show the reaction diagram
additional information
?
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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
p-stomatin PH1511 + H2O
?
show the reaction diagram
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the N-terminal region of PH1510 (residues 16-236, i.e. 1510-N) is a serine protease with a catalytic Ser-Lys dyad (Ser97 and Lys138) and specifically cleaves the C-terminal hydrophobic region of the p-stomatin PH1511
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?
p-stomatin PH1511p + H2O
?
show the reaction diagram
O59179
the N-terminal region of PH1510p is a serine protease and specifically cleaves the C-terminal hydrophobic region of the p-stomatin PH1511p
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?
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
3,4-dichloroisocoumarin
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1 mM, 49% inhibition
CaCl2
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0.1 M, 11% activity
dodecyl-beta-D-maltoside
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0.1%, 38% activity
EDTA
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5 mM, 67% inhibition
iodoacetate
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5 mM, 17% inhibition
MgCl2
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0.1 M, 14% residual activity
MnCl2
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0.1 M, 4% activity
NaCl
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0.1 M, 90% loss of activity
Pepstatin
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0.005 mM, 21% inhibition
phenylmethylsulfonyl fluoride
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5 mM, 21% inhibition
SDS
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0.01%, 53% activity. 1% SDS, 1.2% activity
ZnCl2
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0.1 M, 9% activity
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25000
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2 * 25000, dimer of the N-terminal fragment of PH1510 (1510-N, residues 16–236), SDS-PAGE
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
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2 * 25000, dimer of the N-terminal fragment of PH1510 (1510-N, residues 16–236), SDS-PAGE
hexamer
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molecular modeling of hexameric oligonucleotide binding domains of enzyme C-terminal region of enzyme PH1510 , inter-domain interactions and domain structure, overview
multimer
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the structure of C-terminal soluble enzyme domain 1510-C, residues 371-441 of PH1510, has a compact five-stranded beta-barrel fold known as an oligosaccharide/oligonucleotide-binding fold (OB-fold). According to crystal packing, 1510-C can assemble into multimers based on a dimer as a basic unit. C-terminal soluble enzyme domain 1510-C also forms a large cylinder-like structure composed of 24 subunits or a large triangular prism-like structure composed of 12 subunits
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
crystal structure of 1510-N (N-terminal region of open reading frame, PH1510 (residues 16–236)) in dimeric form
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enzyme mutant K138A in complex with a peptide substrate, an enzyme dimer bound to one peptide, X-ray diffraction structure determination and analysis at 2.25 A resolution
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hanging-drop vapor diffusion method, crystallization of the catalytically inactive mutants S97A and K138A of 1510-N in complex with three kinds of substrate peptides of p-stomatin (234NVIVLMLPME243, 232KSNVIVLML240 and 238LMLPMEMLK246), structural analysis
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purified recombinant C-terminal soluble domain 1510-C of STOPP PH1510, sitting drop vapor diffusion method, mixing of 0.0005 ml of 5.7 mg/ml protein in 50 mM Tris-HCl, pH 7.5, and 50 mM NaCl with 0.0005 ml of reservoir solution containing 20% v/v Jeffamine M-600 and 0.1 M HEPES-NaOH, pH 7.5, 20°C, X-ray diffraction structure determination and analysis at 2.4 A resolution, molecular replacement, for modelling the structure with PDB ID 2EXD is used method
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purification of the N-terminal fragment of PH1510 (1510-N, residues 16–236) expressed in Escherichia coli
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recombinant His-tagged full-length enzyme from Escherichia coli strain BL21-CodonPlus by nickel affinity and anion exchange chromatography, followed by dialysis
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
gene maps of the Pyrococcus horikoshii genome containing two sets of STOPP/stomatin gene pairs, i.e. PH1510 (long-STOPP)/PH1511 (p-stomatin) and PH0471 (short-STOPP)/PH0470 (p-stomatin). Recombinant expression of His-tagged full-length enzyme in Escherichia coli strain BL21-CodonPlus
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the N-terminal region of PH1510 (1510-N, residues 16–236) is expressed in Escherichia coli
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D168A
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mutant enzyme shows 3.2% activity with casein compared with the activity of the wild-type enzyme. The mutant enzyme shows a 25000 Da band instead of 45000 Da band
S97A
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mutant enzyme shows 0.08% activity with casein compared with the activity of the wild-type enzyme
T62A
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mutant enzyme shows 5.5% activity with casein compared with the activity of the wild-type enzyme