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Information on EC 3.4.21.62 - Subtilisin
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3.4.21.62 SubtilisinSpecify your search results
Word Map
- 3.4.21.62
- proteases
- hydrolysates
- chymotrypsin
- prion
- papain
- elastase
- prpsc
- aeruginosa
- sds
- scrapie
-
convertase
- alpha-chymotrypsin
- proprotein
- pronase
- casein
- dnase
-
protease-resistant
- pcsk9
- pmsf
-
proteinaceous
- propeptide
-
bacteriocins
-
spongiform
- licheniformis
-
ultrafiltration
- thermolysin
- alpha-amylase
-
shiga
-
formalin-fixed
-
phenylmethylsulfonyl
- creutzfeldt-jakob
-
paraffin-embedded
- amyloliquefaciens
-
cytotoxin
- gelatin
-
boil
-
i-converting
- medicine
- pumilus
- album
-
alkaliphilic
- prodomain
-
transesterification
- p-nitroanilide
- biotechnology
- bromelain
- detergent
- furin
-
defatted
-
kazal
- synthesis
- degradation
- analysis
- pancreatin
-
anhydrous
- food industry
- industry
The enzyme appears in viruses and cellular organisms
Reaction Schemes
Hydrolysis of proteins with broad specificity for peptide bonds, and a preference for a large uncharged residue in P1. Hydrolyses peptide amides
Synonyms
AcpII, Ak.1 protease, Alcalase, Alcalase 0.6L, Alcalase 2.5L, ALE1 subtilase, ALK-enzyme, Alkaline mesentericopeptidase, Alkaline protease, alkaline serine protease, 
more
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SYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
AcpII
Ak.1 protease
Alcalase
Alcalase 0.6L
-
-
-
-
Alcalase 2.5L
-
-
-
-
ALK-enzyme
-
-
-
-
Alkaline mesentericopeptidase
-
-
-
-
Alkaline protease
alkaline serine protease
aprE
AtSBT1.9
Bacillopeptidase A
-
-
-
-
Bacillopeptidase B
-
-
-
-
Bacillus gibsonii alkaline protease
Bacillus subtilis alkaline proteinase Bioprase
-
-
-
-
BgAP
Bioprase AL 15
-
-
-
-
Bioprase APL 30
-
-
-
-
cold active subtilisin-like serine proteinase
Colistinase
-
-
-
-
EC 3.4.21.14
EC 3.4.4.16
-
-
formerly
-
Esperase
-
-
-
-
Fe protease
Fe prtS8A
Genenase I
-
-
-
-
IvaP
Kazusase
-
-
-
-
Maxatase
-
-
-
-
Nagarse
-
-
-
-
Opticlean
-
-
-
-
ORF2
Orientase 10B
-
-
-
-
PbSOPT
Peptidase, subtilo-, A
-
-
-
-
phytophase
PIMMS2
Protease S
-
-
-
-
Protease VIII
-
-
-
-
Protease XXVII
-
-
-
-
Proteinase, Bacillus subtilis alkaline
-
-
-
-
Protin A 3L
-
-
-
-
SAP
Savinase
-
-
-
-
Savinase 16.0L
-
-
-
-
Savinase 32.0 L EX
-
-
-
-
Savinase 4.0T
-
-
-
-
Savinase 8.0L
-
-
-
-
SBL
SES7
SOPT
SP 266
-
-
-
-
subtilase
subtilase subfamily 1 member 9
subtilase-like protease
subtilisin
subtilisin 72
subtilisin A
Subtilisin amylosacchariticus
subtilisin BPN’
subtilisin Carlsberg
subtilisin DJ-4
Subtilisin DY
Subtilisin E
subtilisin E-S7
Subtilisin GX
-
-
-
-
subtilisin JB1
Subtilisin Novo
-
-
-
-
subtilisin Pr1-like protease
subtilisin QK
Subtilisin S41
Subtilisin Sendai
-
-
-
-
subtilisin Sph
subtilisin-like ookinete protein
subtilisin-like ookinete protein important for transmission
subtilisin-like protease
subtilisin-like serine protease
Subtilopeptidase
-
-
-
-
Superase
-
-
-
-
thermo-active subtilisin-like serine protease
Thermoase
-
-
-
-
Thermoase PC 10
-
-
-
-
thermostable subtilisin
ThSS45
trans-cinnamoyl-subtilisin
-
acyl-enzyme covalent intermediate of the serine protese subtilisin Carlsberg
V. cholerae-secreted serine protease
VC_0157
vPR
additional information
Alcalase
-
-
-
-
Alkaline protease
-
-
-
-
EC 3.4.21.14
-
-
formerly
-
phytophase
-
extensive homology to other plant subtilisin-like serine protease, phytaspase: a plant aspartate-specific protease
phytophase
-
extensive homology to other plant subtilisin-like serine protease, phytaspase: a plant aspartate-specific protease
Subtilisin amylosacchariticus
-
from Bacillus subtilis var. amylosacchariticus
Subtilisin amylosacchariticus
-
from Bacillus subtilis var. amylosacchariticus
Subtilisin amylosacchariticus
-
from Bacillus subtilis var. amylosacchariticus
subtilisin Carlsberg
-
-
Subtilisin E
-
-
-
-
Subtilisin S41
-
-
-
-
additional information
-
species variants include subtilisin BPN' (also subtilisin B, subtilopeptidase B, subtilopeptidase C, Nagarse, Nagarse proteinase, subtilisin Novo, bacterial proteinase Novo) and subtilisin Carlsberg (subtilisin A, subtilopeptidase A, alcalase Novo), similar enzymes are produced by various Bacillus subtilis strains and other Bacillus species
additional information
-
synonyms of subtilisin Carlsberg: subtilisin, subtilisin A, subtilopeptidase A, alcalase Novo, synonyms of subtilisin BPN': nagarse proteinase, Nagarse, BPN', subtilopeptidase C, synonyms of subtilisin Novo: bacterial proteinase Novo, subtilisin B, subtilopeptidase B
additional information
-
species variants include subtilisin BPN' (also subtilisin B, subtilopeptidase B, subtilopeptidase C, Nagarse, Nagarse proteinase, subtilisin Novo, bacterial proteinase Novo) and subtilisin Carlsberg (subtilisin A, subtilopeptidase A, alcalase Novo), similar enzymes are produced by various Bacillus subtilis strains and other Bacillus species
additional information
-
synonyms of subtilisin Carlsberg: subtilisin, subtilisin A, subtilopeptidase A, alcalase Novo, synonyms of subtilisin BPN': nagarse proteinase, Nagarse, BPN', subtilopeptidase C, synonyms of subtilisin Novo: bacterial proteinase Novo, subtilisin B, subtilopeptidase B
additional information
-
the commercial enzyme subtilisin BPN' from Sigma: kinetic and physical properties indicate that the commercial enzyme is probably subtilisin Carlsberg isolated from Bacillus licheniformis, much work which has been performed on commercial subtilisin BPN' from 1975 up to 1985 must be revaluated
additional information
-
synonyms of subtilisin Carlsberg: subtilisin, subtilisin A, subtilopeptidase A, alcalase Novo, synonyms of subtilisin BPN': nagarse proteinase, Nagarse, BPN', subtilopeptidase C, synonyms of subtilisin Novo: bacterial proteinase Novo, subtilisin B, subtilopeptidase B
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REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Hydrolysis of proteins with broad specificity for peptide bonds, and a preference for a large uncharged residue in P1. Hydrolyses peptide amides
Hydrolysis of proteins with broad specificity for peptide bonds, and a preference for a large uncharged residue in P1. Hydrolyses peptide amides
hydrolysis of proteins with broad specificity for peptide bonds, and a preference for a large uncharged residue in P1. Hydrolyzes peptide amides. Mechanism
-
Hydrolysis of proteins with broad specificity for peptide bonds, and a preference for a large uncharged residue in P1. Hydrolyses peptide amides
hydrolysis of proteins with broad specificity for peptide bonds, and a preference for a large uncharged residue in P1. Hydrolyzes peptide amides. Mechanism
-
Hydrolysis of proteins with broad specificity for peptide bonds, and a preference for a large uncharged residue in P1. Hydrolyses peptide amides
importance of tetrahedral intermediate formation in the catalytic mechanism of the serine protease subtilisin. Substrate binding induces the formation of a strong hydrogen bond or low-barrier hydrogen bond between histidine-57 and aspartate-102 that increases the pK(a) of the active site histidine, allowing it to be an effective general base catalyst for the formation of the tetrahedral intermediate and increasing the effective molarity of the catalytic hydroxyl group of serine-195, catalytic mechanism, overview
-
Hydrolysis of proteins with broad specificity for peptide bonds, and a preference for a large uncharged residue in P1. Hydrolyses peptide amides
-
-
-
-
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ester bond hydrolysis
hydrolysis of peptide bond
-
-
-
-
transesterification
-
-
-
-
transpeptidation
-
-
-
-
ester bond hydrolysis
-
-
-
-
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CAS REGISTRY NUMBER
COMMENTARY
9014-01-1
-
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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
(R,S)-2-benzyl-3-(tert-butylsulfonyl)propionic acid + H2O
(R)-2-benzyl-3-(tert-butylsulfonyl)propionic ethyl ester + (S)-2-benzyl-3-(tert-butylsulfonyl)propionic acid + ethanol
-
-
-
-
?
(R,S)-2-benzyl-3-[[1-methyl-1-((morpholin-4-yl)-carbonyl)ethyl]sulfonyl]propionic acid + H2O
(S)-2-benzyl-3-[[1-methyl-1-((morpholin-4-yl)-carbonyl)ethyl]sulfonyl]propionic acid + (R)-2-benzyl-3-[[1-methyl-1-((morpholin-4-yl)-carbonyl)ethyl]sulfonyl]propionic acid-ethyl ester + ethanol
-
-
-
-
?
(R/S)-(2-methylpropyl)butanedioic acid diethyl ester + H2O
(S)-(2-methylpropyl)butanedioic acid diethyl ester + (R)-(2-methylpropyl)butanedioic acid 4-ethyl ester + ethanol
-
-
-
-
?
2 rac-1-(4-chlorophenyl)ethan-1-ol + vinyl butyrate
(1S)-1-(4-chlorophenyl)ethyl butyrate + (1R)-1-(4-chlorophenyl)ethan-1-ol + ?
-
-
-
?
2 rac-1-(4-methoxyphenyl)ethan-1-ol + vinyl butyrate
(1S)-1-(4-methoxyphenyl)ethyl butyrate + (1R)-1-(4-methoxyphenyl)ethan-1-ol + ?
substrate only of mutant G165L/M221F, not of the wild-type enzyme
-
-
?
2 rac-1-(4-methylphenyl)ethan-1-ol + vinyl butyrate
(1S)-1-(4-methylphenyl)ethyl butyrate + (1R)-1-(4-methylphenyl)ethan-1-ol + ?
substrate only of mutant G165L/M221F, not of the wild-type enzyme
-
-
?
2 rac-1-phenylbutan-1-ol + vinyl butyrate
(S)-1-phenylbutyl butyrate + (R)-1-phenylbutan-1-ol + ?
substrate only of mutant G165L/M221F, not of the wild-type enzyme
-
-
?
2 rac-1-phenylethanol + vinyl butyrate
(S)-1-phenylethyl butyrate + (R)-1-phenylethanol + ?
-
-
-
?
2 rac-1-phenylheptan-1-ol + vinyl butyrate
(S)-1-phenylheptyl butyrate + (R)-1-phenylheptan-1-ol + ?
substrate only of mutant G165L/M221F, not of the wild-type enzyme
-
-
?
2 rac-1-phenylpentan-1-ol + vinyl butyrate
(S)-1-phenylpentyl butyrate + (R)-1-phenylpentan-1-ol + ?
substrate only of mutant G165L/M221F, not of the wild-type enzyme
-
-
?
2 rac-1-phenylpropan-1-ol + vinyl butyrate
(S)-1-phenylpropyl butyrate + (R)-1-phenylpropan-1-ol + ?
substrate only of mutant G165L/M221F, not of the wild-type enzyme
-
-
?
2 rac-N-tert-butoxycarbonylphenylalanine ethyl thioester + 1-phenylmethanamine + tert-butanol
N-benzyl-Nalpha-(tert-butoxycarbonyl)-L-phenylalaninamide + (R)-N-tert-butoxycarbonylphenylalanine ethyl thioester + ?
-
continuous-flow cascade reactor system for subtilisin A-catalyzed dynamic kinetic resolution (DKR). The continuous-mode DKR of the racemic thioester in a serial cascade system of six biocatalyst-filled columns at 50°C for KR and five grafted silica gel-filled columns at 150°C for racemization results in the formation of the (S)-benzylamide in 79% conversion, 8.17 g/l/h volumetric productivity and 98% enantiomeric excess. Method evaluation, overview
-
-
?
67 kDa gamma-glutamyl transpeptidase + H2O
30 kDa gamma-glutamyl transpeptidase + ?
-
proteolytic digestion of 67 kDa gamma-glutamyl transpeptidase from Bacillus licheniformis ER-15 by subtilisin to the 30 kDa form, which in turn remains associated with subtilisin as a heterodimeric protein
-
-
?
acid casein + H2O
?
-
purified caseins from animal's milk from cow, sheep, goat and water buffalo used as substrate for subtilisin, cow acid casein is the best substrate
-
?
Ala-Ala-Pro-Leu-4-nitroanilide + H2O
Ala-Ala-Pro-Leu + 4-nitroaniline
low activity
-
-
?
Ala-Ala-Pro-Lys-4-nitroanilide + H2O
Ala-Ala-Pro-Lys + 4-nitroaniline
low activity
-
-
?
Ala-Ala-Pro-Met-4-nitroanilide + H2O
Ala-Ala-Pro-Met + 4-nitroaniline
low activity
-
-
?
Ala-Ala-Pro-Nle-4-nitroanilide + H2O
Ala-Ala-Pro-Nle + 4-nitroaniline
low activity
-
-
?
Ala-Ala-Pro-Phe-4-nitroanilide + H2O
Ala-Ala-Pro-Phe + 4-nitroaniline
low activity
-
-
?
Ala-Ala-Val-Ala-4-nitroanilide + H2O
Ala-Ala-Val-Ala + 4-nitroaniline
low activity
-
-
?
benzyloxycarbonyl-Gly-4-nitroanilide + H2O
benzyloxycarbonyl-Gly + 4-nitroaniline
-
-
-
-
?
benzyloxycarbonyl-L-Ala ethyl ester + butanol
benzyloxycarbonyl-L-Ala butyl ester + ethanol
-
in isooctane, preferential use of L-enantiomer. Comparison with enantioselectivity in water and DMSO
-
-
?
benzyloxycarbonyl-L-Ala-L-Ala-L-Leu-4-nitroanilide + H2O
benzyloxycarbonyl-L-Ala-L-Ala-L-Leu + 4-nitroaniline
benzyloxycarbonyl-L-Asp methyl ester + serine amide
benzyloxycarbonyl-L-Asp-L-Ser-NH2 + methanol
-
-
-
-
?
Carboxybenzoyl-Gly-Ala-NH2 + H2O
Carboxybenzoyl-Gly-Ala + NH3
-
subtilisin BPN'
subtilisin BPN'
?
Carboxybenzoyl-Gly-Leu-NH2 + H2O
Carboxybenzoyl-Gly-Leu + NH3
-
subtilisin BPN'
subtilisin BPN'
?
Carboxybenzoyl-Gly-Pro-Leu-Gly-Pro-OH + H2O
Carboxybenzoyl-Gly-Pro-Leu + Gly-Pro-OH
-
subtilisin BPN'
subtilisin BPN'
?
D-Val-Leu-Lys p-nitroanilide + H2O
D-Val-Leu-Lys + p-nitroaniline
-
-
-
?
eglin-c + H2O
?
-
proteinase inhibitor, subtilisin DY hydrolyses the peptide bond between residues 45 and 46 in the reactive centre of eglin-c
-
?
ethyl 2-(4-ethylphenoxy)propionate + butanol
butyl 2-(4-ethylphenoxy)propanoate + propanol
-
in isooctane, preferential use of S-enantiomer. Comparison with enantioselectivity in water and DMSO
-
-
?
glutaryl-Ala-Ala-Pro-Leu-p-nitroanilide + H2O
glutaryl-Ala-Ala-Pro-Leu + p-nitroaniline
relative hydrolysis rate is 22%
-
-
?
Human fibrinogen + H2O
?
-
fibrinolytic activity is determined using the fibrin plate method
-
-
?
IvaP I9 domain + H2O
?
a temporary inhibitor and substrate of purified IvaP
-
-
?
kappa casein + H2O
?
-
purified caseins from animal's milk from cow, sheep, goat and water buffalo used as substrate for subtilisin
-
?
L-phenylalanine-isopropylester + H2O
L-phenylalanine + isopropanol
-
-
-
-
?
lactate dehydrogenase + H2O
?
poor substrate in native form, but can be digested in heat-denatured form
-
-
?
malate dehydrogenase + H2O
?
poor substrate in native form, but can be digested in heat-denatured form
-
-
?
methoxysuccinyl-Ala-Ile-Pro-Met-p-nitroanilide + H2O
methoxysuccinyl-Ala-Ile-Pro-Met + p-nitroaniline
is the most favorable substrate, relative hydrolysis rate is 100%
-
-
?
methyl mandelate + butanol
butyl mandelate + methanol
-
in isooctane, preferential use of S-enantiomer. Comparison with enantioselectivity in water and DMSO
-
-
?
N-acetyl-L-phenylalanine ethyl ester + 1-butanol
?
-
transesterification catalyzed by poly(ethylene glycol)-modified subtilisin
-
-
?
N-methoxysuccinyl-Ala-Ala-Pro-Val-4-nitroanilide + H2O
N-methoxysuccinyl-Ala-Ala-Pro-Val + 4-nitroaniline
-
-
-
-
?
N-methoxysuccinyl-Ala-Ala-Pro-Val-p-nitroanilide + H2O
N-methoxysuccinyl-Ala-Ala-Pro-Val + p-nitroaniline
N-Succ-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
N-Succ-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
-
?
N-succinyl-Ala-Ala-Pro-Leu-4-nitroanilide + H2O
N-succinyl-Ala-Ala-Pro-Leu + 4-nitroaniline
-
-
-
?
N-succinyl-Ala-Ala-Pro-Met-p-nitroanilide + H2O
N-succinyl-Ala-Ala-Pro-Met + p-nitroaniline
relative hydrolysis rate is 39%
-
-
?
N-succinyl-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
N-succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide + H2O
N-succinyl-Ala-Ala-Pro-Phe + p-nitroaniline
N-succinyl-Ala-Ala-Val-Ala-4-nitroanilide + H2O
N-succinyl-Ala-Ala-Val-Ala + 4-nitroaniline
-
-
-
?
Nalpha-benzoyl-DL-Arg-4-nitroanilide + H2O
Nalpha-benzoyl-DL-Arg + 4-nitroaniline
-
-
-
-
?
oxidized insulin chain B + H2O
?
-
is cleaved by subtilisin-like serine protease at multiple sites, preferably at the C-termini of the hydrophobic residues, such as Tyr, Phe, Leu, Val and Ala. This peptide is also cleaved by subtilisin at the C-termini of the hydrophobic residues, but at more specific sites
-
-
?
p-nitrophenyl butyrate + H2O
p-nitrophenol
-
catalyzed by native subtilisin
-
-
?
Phe-Ala-Ala-Phe-4-nitroanilide + H2O
Phe-Ala-Ala-Phe + 4-nitroaniline
highly preferred substrate
-
-
?
porcine brain tubulin + H2O
peptides
-
MALDI-mass spectrum of the carboxy-terminal peptides released by subtilisin treatment. Polyglutamylated peptides from the beta subunit and from both the tyrosinated and detyrosinated forms of the alpha subunit are observed. The fragments removed by subtilisin are in the range of 1.6 kDa. Post-translational modifications can be directly identified in the mixture of peptides resulting from limited subtilisin treatment
-
-
?
succinyl-AAPF-p-nitroanilide + H2O
succinyl-alanyl-alanyl propyl-phenylalanine
-
-
-
?
succinyl-Ala-Ala-Ala-Ala-Ala-p-nitroanilide + H2O
succinyl-AAAAA + p-nitroaniline
-
-
-
?
succinyl-Ala-Ala-Ala-p-nitroanilide + H2O
succinyl-AAA + p-nitroaniline
-
-
-
?
succinyl-Ala-Ala-p-nitroanilide + H2O
succinyl-Ala-Ala + p-nitroaniline
-
-
-
?
succinyl-Ala-Ala-Phe-4-nitroanilide + H2O
succinyl-Ala-Ala-Phe + 4-nitroaniline
-
-
-
-
?
succinyl-Ala-Ala-Pro-Ala-p-nitroanilide + H2O
succinyl-AAPA + p-nitroaniline
-
-
-
?
succinyl-Ala-Ala-Pro-Leu-p-nitroanilide + H2O
succinyl-AAPL + p-nitroaniline
-
-
-
?
succinyl-Ala-Ala-Pro-Phe-7-amido-4-methylcoumarin + H2O
succinyl-Ala-Ala-Pro-Phe + 7-amino-4-methylcoumarin
-
-
-
-
?
succinyl-Ala-Ala-Pro-Phe-p-nitroanilide + H2O
succinyl-Ala-Ala-Pro-Phe + p-nitroaniline
-
-
-
-
?
succinyl-L-Ala-L-Ala-L-Phe-7-amido-4-methylcoumarin + H2O
succinyl-L-Ala-L-Ala-L-Phe + 7-amino-4-methylcoumarin
-
-
-
-
?
succinyl-L-Ala-L-Ala-L-Pro-Gly-4-nitroanilide + H2O
succinyl-L-Ala-L-Ala-L-Pro-Gly + 4-nitroaniline
-
-
-
-
?
succinyl-L-Ala-L-Ala-L-Pro-L-Ala-4-nitroanilide + H2O
succinyl-L-Ala-L-Ala-L-Pro-L-Ala + 4-nitroaniline
-
-
-
-
?
succinyl-L-Ala-L-Ala-L-Pro-L-Lys-4-nitroanilide + H2O
succinyl-L-Ala-L-Ala-L-Pro-L-Lys + 4-nitroaniline
-
-
-
-
?
succinyl-L-Ala-L-Ala-L-Pro-L-Met-4-nitroanilide + H2O
succinyl-L-Ala-L-Ala-L-Pro-L-Met + 4-nitroaniline
-
-
-
-
?
succinyl-L-Ala-L-Ala-L-Pro-L-Phe-4-methyl-coumaryl-7-amide + H2O
succinyl-L-Ala-L-Ala-L-Pro-L-Phe + 7-amino-4-methylcoumarin
-
-
-
?
succinyl-L-Ala-L-Ala-L-Pro-L-Phe-4-nitroanilide + H2O
succinyl-L-Ala-L-Ala-L-Pro-L-Phe + 4-nitroaniline
-
-
-
-
?
succinyl-L-Ala-L-Ala-L-Pro-L-Tyr-4-nitroanilide + H2O
succinyl-L-Ala-L-Ala-L-Pro-L-Tyr + 4-nitroaniline
-
-
-
-
?
succinyl-L-Asp-L-Val-L-Arg-L-Ala-L-Phe-7-amido-4-methylcoumarin + H2O
succinyl-L-Asp-L-Val-L-Arg-L-Ala-L-Phe + 7-amino-4-methylcoumarin
-
-
-
-
?
succinyl-Phe-Ala-Ala-Phe-p-nitroanilide + H2O
succinyl-Phe-Ala-Ala-Phe + p-nitroaniline
-
-
-
?
Toluenesulfonyl-Arg methyl ester + H2O
?
-
subtilisin Carlsberg, subtilisin Novo
-
-
?
Z-Ala-Ala-Leu-OMe + Phe-p-nitroanilide
Z-Ala-Ala-Leu-Phe-p-nitroanilide + methanol
-
-
-
-
?
Z-Ala-Phe-OMe + H2O
Z-Ala-Phe-NH2 + methanol
-
enzymatic method for the synthesis of free terminal amides of peptides, by ammonolysis of peptide methyl esters using ammonium carbamate and subtilisin A from Bacillus licheniformis in polar organic solvents with low water content is developed. Enzyme is very stable and active in a mixture of t-BuOH and DMF 82.5:17.5 (v/v), containing 0.2% water. Optimum conditions for Z-Ala-Phe-NH2 synthesis are molar ratio ammonium carbamate to Z-Ala-Phe-OMe 10, in t-BuOH/DMF, 82.5:17.5 (v/v) containing 0.2% (v) water, at 30°C for 21 h with the maximum yield of 87%
-
-
?
Z-Thr-Ala-Thr-OCH3 + Asp-p-nitroanilide
Z-Thr-Ala-Thr-Asp-p-nitroanilide + methanol
-
-
-
-
?
Benzoyl-Arg ethyl ester + H2O
?
-
subtilisin Carlsberg, subtilisin Novo
-
-
?
Benzoyl-Arg ethyl ester + H2O
?
-
subtilisin BPN', subtilisin Amylosacchariticus
-
-
?
benzyloxycarbonyl-L-Ala-L-Ala-L-Leu-4-nitroanilide + H2O
benzyloxycarbonyl-L-Ala-L-Ala-L-Leu + 4-nitroaniline
-
-
-
-
?
benzyloxycarbonyl-L-Ala-L-Ala-L-Leu-4-nitroanilide + H2O
benzyloxycarbonyl-L-Ala-L-Ala-L-Leu + 4-nitroaniline
-
-
-
-
?
Bovine serum albumin + H2O
?
poor substrate in native form, but can be digested in heat-denatured form
-
-
?
casein + H2O
?
38% activity relative to gelatin as substrate. Also purified AcpII-C and AcpII-C-DELTAPA act well on casein
-
-
?
casein + H2O
?
38% activity relative to gelatin as substrate. Also purified AcpII-C and AcpII-C-DELTAPA act well on casein
-
-
?
casein + H2O
?
-
caseinolytic activity is determined using the casein plate method
-
-
?
Collagen + H2O
?
81% activity relative to gelatin as substrate. Also purified AcpII-C and AcpII-C-DELTAPA act well on collagen
-
-
?
Collagen + H2O
?
81% activity relative to gelatin as substrate. Also purified AcpII-C and AcpII-C-DELTAPA act well on collagen
-
-
?
cuticle + H2O
?
-
Pr1-like activity on host and non-host cuticle. In KV01 cultures, at 16 h, similar Pr1-like activity on both aphid and locust cuticle
-
-
?
cuticle + H2O
?
-
Pr1-like activity on host and non-host cuticle. In KV71 cultures, at 16 and 24 h, similar Pr1-like activity on both aphid and locust cuticle. Filtrates from KV71 and KV42 cultures, that contain cocktails of cuticle-degrading enzymes, hydrolyse 3-4fold more protein from aphid than locust cuticle
-
-
?
Elastin + H2O
?
7% activity relative to gelatin as substrate
-
-
?
Fibrinogen + H2O
?
rapid hydrolysis of Aalpha-, Bbeta- and gamma-chains. At very low concentration, no cleavage of gamma-chain
-
-
?
Fibrinogen + H2O
?
rapid hydrolysis of Aalpha-, Bbeta- and gamma-chains. At very low concentration, no cleavage of gamma-chain
-
-
?
Gelatin + H2O
?
best substrate for AcpII. Also purified AcpII-C and AcpII-C-DELTAPA act well on gelatin
-
-
?
Gelatin + H2O
?
best substrate for AcpII. Also purified AcpII-C and AcpII-C-DELTAPA act well on gelatin
-
-
?
Hammarsten casein + H2O
?
-
activation energy for hydrolysis 10.59 kcal per mol
-
-
?
Hammarsten casein + H2O
?
Alkalihalobacillus clausii GMBAE 42
-
activation energy for hydrolysis 10.59 kcal per mol
-
-
?
intelectin + H2O
?
enzyme IvaP is able to cleave purified intelectin, which inhibits intelectin binding to Vibrio cholerae
-
-
?
intelectin + H2O
?
enzyme IvaP is able to cleave purified intelectin, which inhibits intelectin binding to Vibrio cholerae
-
-
?
intelectin + H2O
?
Vibrio cholerae serotype O1 C6706
enzyme IvaP is able to cleave purified intelectin, which inhibits intelectin binding to Vibrio cholerae
-
-
?
intelectin + H2O
?
enzyme IvaP is able to cleave purified intelectin, which inhibits intelectin binding to Vibrio cholerae
-
-
?
Keratin + H2O
?
1.5% activity relative to gelatin as substrate
-
-
?
Keratin + H2O
?
1.5% activity relative to gelatin as substrate
-
-
?
N-Acetyl-Phe methyl ester + H2O
?
-
subtilisin Amylosacchariticus
-
-
?
N-Acetyl-Phe methyl ester + H2O
?
-
subtilisin Carlsberg, subtilisin Novo
-
-
?
N-Acetyl-Trp methyl ester + H2O
?
-
subtilisin Amylosacchariticus
-
-
?
N-Acetyl-Trp methyl ester + H2O
?
-
subtilisin Carlsberg, subtilisin Novo
-
-
?
N-Acetyl-Tyr ethyl ester + H2O
?
-
subtilisin Carlsberg, subtilisin Novo
-
-
?
N-Acetyl-Tyr methyl ester + H2O
?
-
subtilisin Amylosacchariticus
-
-
?
N-Acetyl-Tyr methyl ester + H2O
?
-
subtilisin Carlsberg, subtilisin Novo
-
-
?
N-Acetyl-Val methyl ester + H2O
?
-
subtilisin Carlsberg, subtilisin Novo
-
-
?
N-acetylglucosamine + H2O
?
-
N-acetylglucosamine has little effect on subtilisin production by KV01 (11% of that on cuticle)
-
-
?
N-acetylglucosamine + H2O
?
-
restricted feeding of N-acetylglucosamine (the monomer of chitin) to biomass of KV71 induces protease to 61% of that on cuticle
-
-
?
N-methoxysuccinyl-Ala-Ala-Pro-Val-p-nitroanilide + H2O
N-methoxysuccinyl-Ala-Ala-Pro-Val + p-nitroaniline
-
-
-
?
N-methoxysuccinyl-Ala-Ala-Pro-Val-p-nitroanilide + H2O
N-methoxysuccinyl-Ala-Ala-Pro-Val + p-nitroaniline
-
-
-
?
N-methoxysuccinyl-Ala-Ala-Pro-Val-p-nitroanilide + H2O
N-methoxysuccinyl-Ala-Ala-Pro-Val + p-nitroaniline
-
-
-
?
N-succinyl-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
N-succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
?
N-succinyl-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
N-succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
?
N-succinyl-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
N-succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
?
N-succinyl-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
N-succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
-
?
N-succinyl-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
N-succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
?
N-succinyl-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
N-succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
?
N-succinyl-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
N-succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
Vibrio cholerae serotype O1 C6706
-
-
-
?
N-succinyl-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
N-succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
?
N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide + H2O
N-succinyl-Ala-Ala-Pro-Phe + p-nitroaniline
relative hydrolysis rate is 94%
-
-
?
N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide + H2O
N-succinyl-Ala-Ala-Pro-Phe + p-nitroaniline
-
-
-
-
?
N-succinyl-L-Ala-L-Ala-L-L-Ala-4-nitroanilide + H2O
?
-
3.8% of the activity with N-succinyl-L-Ala-L-Ala-L-Pro-L-Phe-4-nitroanilide
-
-
?
N-succinyl-L-Ala-L-Ala-L-L-Ala-4-nitroanilide + H2O
?
Alkalihalobacillus clausii GMBAE 42
-
3.8% of the activity with N-succinyl-L-Ala-L-Ala-L-Pro-L-Phe-4-nitroanilide
-
-
?
N-succinyl-L-Ala-L-Ala-L-Pro-L-Leu-4-nitroanilide + H2O
?
-
19% of the activity with N-succinyl-L-Ala-L-Ala-L-Pro-L-Phe-4-nitroanilide
-
-
?
N-succinyl-L-Ala-L-Ala-L-Pro-L-Leu-4-nitroanilide + H2O
?
Alkalihalobacillus clausii GMBAE 42
-
19% of the activity with N-succinyl-L-Ala-L-Ala-L-Pro-L-Phe-4-nitroanilide
-
-
?
N-succinyl-L-Ala-L-Ala-L-Pro-L-Leu-4-nitroanilide + H2O
?
-
-
-
-
?
N-succinyl-L-Ala-L-Ala-L-Pro-L-Phe-4-nitroanilide + H2O
?
-
-
-
-
?
N-succinyl-L-Ala-L-Ala-L-Pro-L-Phe-4-nitroanilide + H2O
?
Alkalihalobacillus clausii GMBAE 42
-
-
-
-
?
N-succinyl-L-Ala-L-Ala-L-Pro-L-Phe-4-nitroanilide + H2O
?
-
-
-
-
?
N-succinyl-L-Ala-L-Ala-L-Val-4-nitroanilide + H2O
?
-
2.2% of the activity with N-succinyl-L-Ala-L-Ala-L-Pro-L-Phe-4-nitroanilide
-
-
?
N-succinyl-L-Ala-L-Ala-L-Val-4-nitroanilide + H2O
?
Alkalihalobacillus clausii GMBAE 42
-
2.2% of the activity with N-succinyl-L-Ala-L-Ala-L-Pro-L-Phe-4-nitroanilide
-
-
?
N-tosyl-L-Arg methyl ester + H2O
N-tosyl-L-Arg + methanol
-
-
-
-
?
N-tosyl-L-Arg methyl ester + H2O
N-tosyl-L-Arg + methanol
-
subtilisin Carlsberg, subtilisin Novo, subtilisin Amylosacchariticus
-
-
?
Oxidized insulin B-chain + H2O
?
-
enzyme primarily hydrolyzes Leu15-Tyr16 bond and secondarily Gln4-His5, Ser9-His10, Phe24-Phe25 and Lys29-Ala30
-
-
?
Oxidized insulin B-chain + H2O
?
Bacillus sp. (in: Bacteria) G-825-6
-
enzyme primarily hydrolyzes Leu15-Tyr16 bond and secondarily Gln4-His5, Ser9-His10, Phe24-Phe25 and Lys29-Ala30
-
-
?
Suc-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
Suc-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
-
?
Suc-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
Suc-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
-
?
Suc-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
Suc-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
-
?
Suc-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
Suc-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
-
?
succinyl-AAPF-p-nitroanilide + H2O
succinyl-AAPF + p-nitroaniline
-
-
-
?
succinyl-AAPF-p-nitroanilide + H2O
succinyl-AAPF + p-nitroaniline
-
-
-
?
succinyl-AAPF-p-nitroanilide + H2O
succinyl-AAPF + p-nitroaniline
-
-
-
?
succinyl-AAPF-p-nitroanilide + H2O
succinyl-AAPF + p-nitroaniline
-
-
-
?
succinyl-AAPF-p-nitroanilide + H2O
succinyl-AAPF + p-nitroaniline
-
-
?
succinyl-Ala-Ala-Pro-Phe 4-nitroanilide + H2O
succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
-
?
succinyl-Ala-Ala-Pro-Phe 4-nitroanilide + H2O
succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
-
?
succinyl-Ala-Ala-Pro-Phe 4-nitroanilide + H2O
succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
-
?
succinyl-Ala-Ala-Pro-Phe 4-nitroanilide + H2O
succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
-
?
succinyl-Ala-Ala-Pro-Phe 4-nitroanilide + H2O
succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
-
?
succinyl-Ala-Ala-Pro-Phe 4-nitroanilide + H2O
succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
-
?
succinyl-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
-
?
succinyl-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
-
?
succinyl-Ala-Ala-Pro-Phe-4-nitroanilide + H2O
succinyl-Ala-Ala-Pro-Phe + 4-nitroaniline
-
-
-
-
?
Succinyl-Leu-Leu-Val-Tyr 4-methylcoumarin 7-amide + H2O
?
-
-
-
-
?
Succinyl-Leu-Leu-Val-Tyr 4-methylcoumarin 7-amide + H2O
?
Bacillus sp. (in: Bacteria) G-825-6
-
-
-
-
?
additional information
?
-
substrate specificity, substrate docking simulations, overview. No activity with Ala-Ala-Pro-Glu-4-nitroanilide and Tyr-Val-Ala-Asp-4-nitroanilide. Substrate specificity and the role of stress signals such as divalent metal ions play roles in defining the proteolytic activity of Bacillus clausii intracellular subtilisin protease, molecular basis, overview. The enzyme unfolds under stress conditions. Heat-denatured whole proteins are found to be better substrates for the enzyme than the native forms
-
-
?
additional information
?
-
-
substrate specificity, substrate docking simulations, overview. No activity with Ala-Ala-Pro-Glu-4-nitroanilide and Tyr-Val-Ala-Asp-4-nitroanilide. Substrate specificity and the role of stress signals such as divalent metal ions play roles in defining the proteolytic activity of Bacillus clausii intracellular subtilisin protease, molecular basis, overview. The enzyme unfolds under stress conditions. Heat-denatured whole proteins are found to be better substrates for the enzyme than the native forms
-
-
?
additional information
?
-
N-succinyl-Ala-Ala-Val-Ala-p-nitroanilide and keratin are poor substrates for AcpII, with relative hydrolysis rates of 1-1.5% of the activity toward N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide or gelatin, respectively
-
-
?
additional information
?
-
-
N-succinyl-Ala-Ala-Val-Ala-p-nitroanilide and keratin are poor substrates for AcpII, with relative hydrolysis rates of 1-1.5% of the activity toward N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide or gelatin, respectively
-
-
?
additional information
?
-
N-succinyl-Ala-Ala-Val-Ala-p-nitroanilide and keratin are poor substrates for AcpII, with relative hydrolysis rates of 1-1.5% of the activity toward N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide or gelatin, respectively
-
-
?
additional information
?
-
-
N-succinyl-Ala-Ala-Val-Ala-p-nitroanilide and keratin are poor substrates for AcpII, with relative hydrolysis rates of 1-1.5% of the activity toward N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide or gelatin, respectively
-
-
?
additional information
?
-
-
reactions catalyzed: 1. peptide bond hydrolysis, 2. ester bond hydrolysis, 3. transesterification, 4. transpeptidation
-
-
?
additional information
?
-
-
reactions catalyzed: 1. peptide bond hydrolysis, 2. ester bond hydrolysis, 3. transesterification, 4. transpeptidation
-
-
?
additional information
?
-
-
alteration of substrate specificity by protein engineering
-
-
?
additional information
?
-
-
substrates bind in a less catalytically favorable conformation after the enzyme has been exposed to organic media for several hours
-
-
?
additional information
?
-
besides ester hydrolysis the enzyme also performs ester perhydrolysis reacting an ester and a hydroperpxide, residue Gl165 in the S1 binding pocket of the enzyme is involved. Perhydrolysis reaction analysis and kinetcis, overview
-
-
?
additional information
?
-
-
ionic-surfactant-coated Bacillus licheniformis subtilisin (ISCBLS) is a catalyst for the dynamic kinetic resolution of secondary alcohols. The engineered enzyme ISCBCL displays 9300fold enhanced activity relative to its native counterpart in the transesterificaion of N-acetyl phenylalanine ethyl ester with 1-propanol in hexane and 12800fold enhanced activity in the transesterification of trifluoroethyl butyrate with 1-phenylethanol in tetrahydrofuran. 50 secondary alcohols are evaluated as substrates for kinetic resolution in presence of trifluoroethyl butyrate, product determination and analysis, overview. The engineered enzyme displays enantioselectivity for most of the secondary alcohols tested
-
-
?
additional information
?
-
low catalytic activity of subtilisin Carlsberg (SC) for transacylation reactions with secondary alcohols in organic solvent, tetrahedral intermediates of the model reaction, enzyme with bound (S)-alpha-methylbenzyl butyrate or (R)-alpha-methylbenzyl butyrate, overview. The alkyl chain of the acyl donor, vinyl butyrate, is coordinated into the S1 binding pocket. The alkyl chain of the secondary alcohol is directed into the S1' binding pocket, and the phenyl group is directed towards the solvent. G165 is located in the bottom of the S1 pocket. Enantioselectivities of immobilized wild-type and mutants of subtilisin Carlsberg in the transacylation of racemic 1-phenylethanol and vinyl butyrate in tetrahydrofuran. Molecular modeling
-
-
?
additional information
?
-
-
random mutagenesis to enhance the activity of subtilisin in organic solvents
-
-
?
additional information
?
-
-
under relatively non-aqueous conditions, immobilized subtilisin is able to synthesize phenylacetic acid ethyl ester
-
-
?
additional information
?
-
-
preferentially hydrolyzes the ester bond of Ala, but significant hydrolysis is observed with other aliphatic (Gly, Leu) and aromatic (Tyr) amino acids
-
-
?
additional information
?
-
-
reactions catalyzed: 1. peptide bond hydrolysis, 2. ester bond hydrolysis, 3. transesterification, 4. transpeptidation
-
-
?
additional information
?
-
-
reactions catalyzed: 1. peptide bond hydrolysis, 2. ester bond hydrolysis, 3. transesterification, 4. transpeptidation
-
-
?
additional information
?
-
-
little or no reaction with Glu-Phe-p-nitroanilide, benzyloxycarbonyl-Phe-p-nitroanilide, Suc-Phe-p-nitroanilide, acetyl-Phe-p-nitroanilide, Phe-p-nitroanilide, Gly-p-nitroanilide and Ala-p-nitroanilide at 75°C, and Val-Leu-Lys-p-nitroanilide, benzoyl-Arg-p-nitroanilide, Suc-Ala-Ala-p-nitroanilide, benzoyl-Tyr-p-nitroanilide, benzyloxycarbonyl-Lys-Arg-p-nitroanilide, and benzyloxycarbonyl-Arg-p-nitroanilide at 40°C
-
?
additional information
?
-
-
biosynthesis of subtilisin requires participation of an N-terminal prodomain
-
?
additional information
?
-
-
little or no reaction with Glu-Phe-p-nitroanilide, benzyloxycarbonyl-Phe-p-nitroanilide, Suc-Phe-p-nitroanilide, acetyl-Phe-p-nitroanilide, Phe-p-nitroanilide, Gly-p-nitroanilide and Ala-p-nitroanilide at 75°C, and Val-Leu-Lys-p-nitroanilide, benzoyl-Arg-p-nitroanilide, Suc-Ala-Ala-p-nitroanilide, benzoyl-Tyr-p-nitroanilide, benzyloxycarbonyl-Lys-Arg-p-nitroanilide, and benzyloxycarbonyl-Arg-p-nitroanilide at 40°C
-
?
additional information
?
-
Bacillus sp. (in: Bacteria) GX6644
-
preferentially hydrolyzes the ester bond of Ala, but significant hydrolysis is observed with other aliphatic (Gly, Leu) and aromatic (Tyr) amino acids
-
-
?
additional information
?
-
-
random mutagenesis to enhance the activity of subtilisin in organic solvents
-
-
?
additional information
?
-
-
influence of substrate structure of N-protected peptide nitroanilides of the types: benzyloxycarbonyl-A2-A1 4-nitroanilide, benzyloxycarbonyl-A3-A2-A1 4-nitroanilide, benzyloxycarbonyl-A4-A3-A2-A1 4-nitroanilide, subsite S1 is of broad selectivity: preference for hydrophobic amino acid residues (i.e. leucine and phenylalanine) the beta-branched and the basic amino acid residues cannot interact with the S1 subsite and the hydrolysis of the corresponding peptides occurs exclusively at the A2-A1 bond. If S1/A1 interactions are weak (Ala, norvaline, norleucine) the amino acid residue A1 can interact with subsites S1 and S1' resulting in the hydrolysis of two bonds (A1 4-nitroanilide and A2-A1). The subsite S2 reveals a preference for small amino acid residues
-
-
?
additional information
?
-
-
reactions catalyzed: 1. peptide bond hydrolysis, 2. ester bond hydrolysis, 3. transesterification, 4. transpeptidation
-
-
?
additional information
?
-
the enzyme shows activity towards complex substrates, e.g. skimmed milk
-
-
?
additional information
?
-
the enzyme shows activity towards complex substrates, e.g. skimmed milk
-
-
?
additional information
?
-
performance of a thrombolytic activity assay
-
-
?
additional information
?
-
-
the wild-type enzyme performs autoproteolysis, while the enzyme mutant S221C does not. The cleavage patterns of SES7 is determined by hydrolysates peptide profile, it prefers amino acids Q, T, P, or L at P3, P, V and Q at P2, K, L, N, Q, or F at P1, and F, S, or A at P1'
-
-
?
additional information
?
-
-
influence of substrate structure of N-protected peptide nitroanilides of the types: benzyloxycarbonyl-A2-A1 4-nitroanilide, benzyloxycarbonyl-A3-A2-A1 4-nitroanilide, benzyloxycarbonyl-A4-A3-A2-A1 4-nitroanilide, subsite S1 is of broad selectivity: preference for hydrophobic amino acid residues (i.e. leucine and phenylalanine) the beta-branched and the basic amino acid residues cannot interact with the S1 subsite and the hydrolysis of the corresponding peptides occurs exclusively at the A2-A1 bond. If S1/A1 interactions are weak (Ala, norvaline, norleucine) the amino acid residue A1 can interact with subsites S1 and S1' resulting in the hydrolysis of two bonds (A1 4-nitroanilide and A2-A1). The subsite S2 reveals a preference for small amino acid residues
-
-
?
additional information
?
-
-
random mutagenesis to enhance the activity of subtilisin in organic solvents
-
-
?
additional information
?
-
Bacillus subtilis JN-S7
-
the wild-type enzyme performs autoproteolysis, while the enzyme mutant S221C does not. The cleavage patterns of SES7 is determined by hydrolysates peptide profile, it prefers amino acids Q, T, P, or L at P3, P, V and Q at P2, K, L, N, Q, or F at P1, and F, S, or A at P1'
-
-
?
additional information
?
-
performance of a thrombolytic activity assay
-
-
?
additional information
?
-
-
enzymes prefers cleaving after hydrophobic residues (and in particular P1 leucine)
-
-
?
additional information
?
-
-
the Fusarium equiseti Fe protease has a broad substrate specificity, almost all amino acid residues are accepted at position P1, even though it shows some preference for cleavage at the C-terminal side of asparagine and histidine residues. The S4 subsite of Fe protease favors aspartic acid and threonine, substrate specificity and comprison to other subtilisin and selected fungal subtilisin-like proteases, overview
-
-
?
additional information
?
-
-
the Fe protease is active against beta-casein, while it shows poor activity with cytochrome c and ubiquitin. beta-Casein is fully digested, while cytochrome c and ubiquitin are not. Autoproteolytic degradation (autoproteolysis) of the purified recombinant Fe protease. The Fe protease has a broad substrate specificity: almost all amino acid residues are accepted at positions P1-4 and P1'-P4'. The protease has a slight preference for His and Asn in position P1. There are only two amino acids, Pro and Asp, which are rarely found in position P1. Hydrophobic amino acids are overrepresented in position P2, but Gly, Asp, and the aromatic amino acids Phe and Tyr are rarely found in P2. Gly and Thr are favored in position P3 and Thr and Asp in position P4. Tyrosine is common in the P1' position, unlike Phe which is rarely found in position P1'. Compared to other proteases, the Fe protease has unique cleavage site specificity
-
-
?
additional information
?
-
-
the Fe protease is active against beta-casein, while it shows poor activity with cytochrome c and ubiquitin. beta-Casein is fully digested, while cytochrome c and ubiquitin are not. Autoproteolytic degradation (autoproteolysis) of the purified recombinant Fe protease. The Fe protease has a broad substrate specificity: almost all amino acid residues are accepted at positions P1-4 and P1'-P4'. The protease has a slight preference for His and Asn in position P1. There are only two amino acids, Pro and Asp, which are rarely found in position P1. Hydrophobic amino acids are overrepresented in position P2, but Gly, Asp, and the aromatic amino acids Phe and Tyr are rarely found in P2. Gly and Thr are favored in position P3 and Thr and Asp in position P4. Tyrosine is common in the P1' position, unlike Phe which is rarely found in position P1'. Compared to other proteases, the Fe protease has unique cleavage site specificity
-
-
?
additional information
?
-
-
the Fusarium equiseti Fe protease has a broad substrate specificity, almost all amino acid residues are accepted at position P1, even though it shows some preference for cleavage at the C-terminal side of asparagine and histidine residues. The S4 subsite of Fe protease favors aspartic acid and threonine, substrate specificity and comprison to other subtilisin and selected fungal subtilisin-like proteases, overview
-
-
?
additional information
?
-
-
Sub2 undergoes self-digestion at high concentrations
-
-
?
additional information
?
-
-
enzyme is able to hydrolyze a range of fluorogenic substrates, including AFC derivatives of VEID, VAD, YVAD, IETD, VDVAD and LEHD
-
-
?
additional information
?
-
-
enzyme is able to hydrolyze a range of fluorogenic substrates, including AFC derivatives of VEID, VAD, YVAD, IETD, VDVAD and LEHD
-
-
?
additional information
?
-
degrades protein components of both nematode (Meloidogyne sp.) and insect (Phthorimaea opercullella) eggs
-
-
?
additional information
?
-
-
contains the catalytic triad characteristic of subtilisin family proteinases: motif I (Asp200), motif II (His239), and motif III (Ser568). Has the Gram-positive cell wall anchoring motif (Leu-Pro-X-Thr-Gly) at the carboxy-terminus, which is followed by a hydrophobic domain
-
-
?
additional information
?
-
-
contains the catalytic triad characteristic of subtilisin family proteinases: motif I (Asp200), motif II (His239), and motif III (Ser568). Has the Gram-positive cell wall anchoring motif (Leu-Pro-X-Thr-Gly) at the carboxy-terminus, which is followed by a hydrophobic domain
-
-
?
additional information
?
-
-
autoprocessing of Pro-subtilisin-like serine protease
-
-
?
additional information
?
-
Ser361 contributes to IvaP autoprocessing in stationary-phase cultures
-
-
?
additional information
?
-
enzyme IvaP performs autoprocessing involving residue Ser361. Identification of the N-terminal enzyme IvaP cleavage sequence AAQDNV
-
-
?
additional information
?
-
Ser361 contributes to IvaP autoprocessing in stationary-phase cultures
-
-
?
additional information
?
-
enzyme IvaP performs autoprocessing involving residue Ser361. Identification of the N-terminal enzyme IvaP cleavage sequence AAQDNV
-
-
?
additional information
?
-
Vibrio cholerae serotype O1 C6706
Ser361 contributes to IvaP autoprocessing in stationary-phase cultures
-
-
?
additional information
?
-
Vibrio cholerae serotype O1 C6706
enzyme IvaP performs autoprocessing involving residue Ser361. Identification of the N-terminal enzyme IvaP cleavage sequence AAQDNV
-
-
?
additional information
?
-
Ser361 contributes to IvaP autoprocessing in stationary-phase cultures
-
-
?
additional information
?
-
enzyme IvaP performs autoprocessing involving residue Ser361. Identification of the N-terminal enzyme IvaP cleavage sequence AAQDNV
-
-
?
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NATURAL SUBSTRATE
NATURAL 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
IvaP I9 domain + H2O
?
a temporary inhibitor and substrate of purified IvaP
-
-
?
intelectin + H2O
?
enzyme IvaP is able to cleave purified intelectin, which inhibits intelectin binding to Vibrio cholerae
-
-
?
intelectin + H2O
?
enzyme IvaP is able to cleave purified intelectin, which inhibits intelectin binding to Vibrio cholerae
-
-
?
intelectin + H2O
?
Vibrio cholerae serotype O1 C6706
enzyme IvaP is able to cleave purified intelectin, which inhibits intelectin binding to Vibrio cholerae
-
-
?
intelectin + H2O
?
enzyme IvaP is able to cleave purified intelectin, which inhibits intelectin binding to Vibrio cholerae
-
-
?
additional information
?
-
-
biosynthesis of subtilisin requires participation of an N-terminal prodomain
-
?
additional information
?
-
Ser361 contributes to IvaP autoprocessing in stationary-phase cultures
-
-
?
additional information
?
-
Ser361 contributes to IvaP autoprocessing in stationary-phase cultures
-
-
?
additional information
?
-
Vibrio cholerae serotype O1 C6706
Ser361 contributes to IvaP autoprocessing in stationary-phase cultures
-
-
?
additional information
?
-
Ser361 contributes to IvaP autoprocessing in stationary-phase cultures
-
-
?
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Ca2+
Cl-
treatment with CsCl increases the activity several fold. Two Cl- ions are close to the mouth of the active site cleft, where they may affect catalysis
Cs+
treatment with CsCl increases the activity several fold. Two Cs+ ions are close to the mouth of the active site cleft, where they may affect catalysis
Cu2+
KCl
increases enzymatic activity somewhat, while choline-Cl has a larger effect
Mg2+
Sr2+
-
increases thermostability, but to a significantly lower degree than Ca2+
additional information
Ca2+
is important for ISP activity through structural changes. The looped turn constituted by residues Ala180-Pro197 binds Ca2+. Removal of Ca2+ at sites close to the dimer interface and the S1 pocket are involved enzyme inhhibition by EDTA
Ca2+
enzyme activity and/or stability depends on the presence of divalent cations, probably Ca2+ ions, near the surface of the enzyme
Ca2+
-
stabilizing mutations are usually calcium-dependent in their stabilizing effect
Ca2+
-
in the presence of calcium the half-life at 55°C and 60°C are 3.6fold and 3.48fold higher for the native enzyme compared to that in the absence of added calcium. In the presence of 10 mM calcium the half-life of the enzyme at 60°C increases by 6.06fold, 5.20fold and 2.92fold when coupled with oxidized sucrose polymers OSP400, OSP70 and polyglutaraldehyde, respectively
Ca2+
-
as found in other Bacillus subtilisins, the structure of wild-type Savinase contains 2 calcium ion-binding sites
Ca2+
-
bound calcium ions play a key role in protecting against autolysis and thermal denaturation
Ca2+
bound calcium ions play a key role in protecting against autolysis and thermal denaturation
Ca2+
importance of calcium in the medium after enzyme induction, both for stability of the proteinase and cell health. The two calcium binding sites have apparent binding constants in the mM range. Binding of calcium to the weaker of those two sites only affects resistance of the enzyme against irreversible thermal inactivation. Kinetics
Ca2+
-
required, the presence of a low-affinity Ca2+ binding site formed by the backbone carbonyls cannot be excluded based on the amino acid sequence comparisons
Ca2+
-
required, the enzyme sequence contains a low-affinity Ca2+ binding site
Ca2+
calcium-loaded state of five ions bound to each of the two subtilisin molecules. Three of the binding sites have two side chains of an acidic residue coordinating the calcium ion, whereas the other two binding sites have either a main-chain carbonyl, or only one acidic residue side chain coordinating the calcium ion
Ca2+
-
calcium ion binds weakly to the Ca-7 site in the unautoprocessed form, but is trapped upon autoprocessing. The Ca-7 site is required to promote the autoprocessing reaction by stabilizing the autoprocessed form, in which the new N-terminus of the mature domain is structurally disordered
Ca2+
-
six calcium ions bind to pro-S255A at the loop regions. The at least six Ca2+ ions bind to pro-S255A too tightly to be removed by dialysis
Ca2+
-
Ca2+-binding loop is required for folding of subtilisin but does not seriously contribute to the stabilization of subtilisin in a native structure
Ca2+
the enzyme contains 7 Ca2+ ions, 4 of which are responsible for folding requirement of Ca2+ ions for the hyperthermostability of Tk-subtilisin, the Ca1, Ca6, and Ca7 ions, especially the Ca1 ion, contribute to the hyperthermostabilization of Tk-subtilisin
additional information
metal ions are not involved in the subtilisin catalytic mechanism but are an important structural element
additional information
-
metal ions are not involved in the subtilisin catalytic mechanism but are an important structural element
additional information
-
metal ion sites are not formed until late in folding and do not influence the kinetics of folding to the intermediate complex
additional information
enzymatic activity of subtilisin crystals in acetonitrile is sensitive to the type of counterions present before transfer to the organic solvent. Larger cations increase the enzyme activity
additional information
-
strain DJ-4, not significantly affected by Ca2+, Co2+, Mg2+, or Mn2+
additional information
-
unlike subtilisin, subtilisin-like serine protease does not require Ca2+ for folding
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INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
(dimethylamino)-1-naphthalenesulfonyl fluoride
-
100% inhibition of enzyme activity with N-trans cinnamoyl imidazole as substrate, during prolonged exposure to organic solvents the active-site fluorescent label inhibitor adopts a different binding conformation
1,10-phenanthroline
at 37°C and pH of 7.5, 0.1 mM inhibits prosubtilisin JB1 by 40%
2,2'-[[(1-[[(benzyloxy)carbonyl]amino]-2-phenylethyl)phosphoryl]bis(oxybenzene-4,1-diyl)]diacetic acid
-
-
2,2'-[[(1-[[(benzyloxy)carbonyl]amino]-3-methylbutyl)phosphoryl]bis(oxybenzene-4,1-diyl)]diacetic acid
-
-
2,2'-[[(1-[[1-(tert-butoxycarbonyl)-L-prolyl]amino]-3-methylbutyl)phosphoryl]bis(oxybenzene-3,1-diyl)]diacetic acid
-
-
5-Dimethylaminonaphthalene-1-sulfonate
-
circular-dichroism and fluorescence studies of 5-dimethylaminonaphthalene-1-sulfonyl derivative of subtilisin indicate a closely similar structure to that of native subtilisin
acetonitril
high concentrations of acetonitrile significantly decrease the enzyme activity by over 60%, most likely due to disrupting the enzyme structural integrity
alpha2-Macroglobulin
-
inhibits alkaline proteolytic activity of purified Arp by 80%
-
Benzyloxycarbonyl-(Ala)n-PheCH2Cl
-
benzyloxycarbonyl-(Ala)2-PheCH2Cl is the best inhibitor
Benzyloxycarbonyl-L-phenylalanylbromomethane
-
reactivity is about an order of magnitude less than that of subtilisins BPN' and Carlsberg
bis(2,3,5-trimethylphenyl) (1-[[(benzyloxy)carbonyl]amino]-3-methylbutyl)phosphonate
-
-
bis(2,3-dimethylphenyl) (1-[[(benzyloxy)carbonyl]amino]-2-phenylethyl)phosphonate
-
-
bis(2,3-dimethylphenyl) (1-[[(benzyloxy)carbonyl]amino]-3-methylbutyl)phosphonate
-
-
bis(2,5-dimethylphenyl) (1-[[(benzyloxy)carbonyl]amino]-3-methylbutyl)phosphonate
-
-
bis(2-methylphenyl) (1-[[(benzyloxy)carbonyl]amino]-2-phenylethyl)phosphonate
-
-
bis(2-methylphenyl) (1-[[(benzyloxy)carbonyl]amino]-3-methylbutyl)phosphonate
-
-
bis(3,4,5-trimethylphenyl) (1-[[(benzyloxy)carbonyl]amino]-2-phenylethyl)phosphonate
-
-
bis(3,4,5-trimethylphenyl) (1-[[(benzyloxy)carbonyl]amino]-3-methylbutyl)phosphonate
-
-
bis(3,4-dimethylphenyl) (1-[[(benzyloxy)carbonyl]amino]-2-phenylethyl)phosphonate
-
-
bis(3,4-dimethylphenyl) (1-[[(benzyloxy)carbonyl]amino]-3-methylbutyl)phosphonate
-
-
bis(3-chlorophenyl) (1-[[(benzyloxy)carbonyl]amino]-2-phenylethyl)phosphonate
-
-
bis(3-chlorophenyl) (1-[[(benzyloxy)carbonyl]amino]-3-methylbutyl)phosphonate
-
-
bis(3-methoxyphenyl) (1-[[(benzyloxy)carbonyl]amino]-3-methylbutyl)phosphonate
-
-
bis(4-chlorophenyl) (1-[[(benzyloxy)carbonyl]amino]-2-phenylethyl)phosphonate
-
-
bis(4-chlorophenyl) (1-[[(benzyloxy)carbonyl]amino]-3-methylbutyl)phosphonate
-
-
bis(4-ethylphenyl) (1-[[(benzyloxy)carbonyl]amino]-2-phenylethyl)phosphonate
-
-
bis(4-ethylphenyl) (1-[[(benzyloxy)carbonyl]amino]-3-methylbutyl)phosphonate
-
-
bis(4-methoxyphenyl) (1-[[(benzyloxy)carbonyl]amino]-2-phenylethyl)phosphonate
-
-
bis(4-methylphenyl) (1-[[(benzyloxy)carbonyl]amino]-3-methylbutyl)phosphonate
-
-
bis(4-tert-butylphenyl) (1-[[(benzyloxy)carbonyl]amino]-2-phenylethyl)phosphonate
-
-
bis(4-tert-butylphenyl) (1-[[(benzyloxy)carbonyl]amino]-3-methylbutyl)phosphonate
-
-
bis[4-(methylsulfonyl)phenyl] (1-[[(benzyloxy)carbonyl]amino]-2-phenylethyl)phosphonate
-
-
bis[4-(propan-2-yl)phenyl] (1-[[(benzyloxy)carbonyl]amino]-2-phenylethyl)phosphonate
-
-
bis[4-(propan-2-yl)phenyl] (1-[[(benzyloxy)carbonyl]amino]-3-methylbutyl)phosphonate
-
-
bis[4-(sulfanylmethyl)phenyl] (1-[[(benzyloxy)carbonyl]amino]-2-phenylethyl)phosphonate
-
-
bis[4-(sulfanylmethyl)phenyl] (1-[[(benzyloxy)carbonyl]amino]-3-methylbutyl)phosphonate
-
-
Brij 35
at 37°C and pH of 7.5, 0.01% reduces prosubtilisin JB1 relative activity to 30% and 0.05% reduces prosubtilisin JB1 relative activity to 70%
Ca2+
at 37°C and pH of 7.5, 1 mM reduces prosubtilisin JB1 relative activity to 57% and 5 mM reduces prosubtilisin JB1 relative activity to 45%
Chymotrypsin I inhibitor from potato
-
subtilisin Carlsberg, subtilisin BPN'
-
Co2+
at 37°C and pH of 7.5, 1 mM reduces prosubtilisin JB1 relative activity to 14% and 5 mM reduces prosubtilisin JB1 relative activity to 24%
CrSPI-1
-
Kazal-type inhibitor from the hepatopancreas of the Carcinoscorpius rotundicauda, potently inhibits subtilisin
-
diphenyl (1-[[(benzyloxy)carbonyl]amino]-2-phenylethyl)phosphonate
-
-
diphenyl (1-[[(benzyloxy)carbonyl]amino]-3-methylbutyl)phosphonate
-
-
E-64c
at 37°C and pH of 7.5, 0.1 mM inhibits prosubtilisin JB1 by 25%
EGTA
at 37°C and pH of 7.5, 0.1 mM inhibits prosubtilisin JB1 by 34%
EPI1a
-
four-cysteine atypical Kazal-domain of protease inhibitor EPI1 from Phytophthora infestans. 80% inhibition at 0.00015 mM
-
EPI1b
-
typical Kazal-domain of protease inhibitor EPI1 from Phytophthora infestans, little inhibitory effect
-
fungal protease inhibitor F
-
specific inhibitor toward subtilisin-type protease. P1 residue most signficantly affects inhibitory specificity. Mutant T29M has stronger subtilisin-inhibitory activity than the wild-type, mutants T29E and T29R are relatively weaker inhibitors. Inhibitory activities of mutants T29F and T29L are as strong as that of the wild-type
-
guanidine hydrochloride
-
at 2 M inhibits subtilisin-like serine protease by 35% and at 4 M almost completely, whereas it has no inhibitory effect on subtilisin
H2O2
-
the enzyme activity is reduced to 68% after a 20-min incubation with 0.33% hydrogen peroxide at 25°C in Tris buffer, substrate suc-Ala-Ala-Pro-Phe-4-nitroanilide, recombinant enzyme
Hg+
at 37°C and pH of 7.5, 1 mM reduces prosubtilisin JB1 relative activity to 33% and 5 mM reduces prosubtilisin JB1 relative activity to 22%
Inhibitor from seeds of Setaria italica
-
purification and characterization
-
IvaP I9 domain
the N-terminal propeptide of enzyme IvaP, the IvaP I9 domain, can temporarily inhibit, and be cleaved by, the purified enzyme
-
K+
at 37°C and pH of 7.5, with 1 mM results in an almost complete reduction of prosubtilisin JB1 activity, 5 mM reduces prosubtilisin JB1 relative activity to 53%
Mg2+
at 37°C and pH of 7.5, 1 mM reduces prosubtilisin JB1 relative activity to 79% and 5 mM reduces prosubtilisin JB1 relative activity to 48%
N-((tert-Butoxycarbonyl)alanylprolylphenylalanyl)-O-benzoylhydroxylamine
-
-
N-(tert-butoxycarbonyl)-L-valyl-N-(1-[bis[4-(sulfanylmethyl)phenoxy]phosphoryl]-2-phenylethyl)-L-prolinamide
-
-
N-(tert-butoxycarbonyl)-L-valyl-N-[1-(diphenoxyphosphoryl)-2-phenylethyl]-L-prolinamide
-
-
N-(tert-butoxycarbonyl)-L-valyl-N-[1-[bis(3,4,5-trimethylphenoxy)phosphoryl]-3-methylbutyl]-L-prolinamide
-
-
N-(tert-butoxycarbonyl)-L-valyl-N-[1-[bis(4-methoxyphenoxy)phosphoryl]-3-methylbutyl]-L-prolinamide
-
-
N-(tert-butoxycarbonyl)-L-valyl-N-[1-[bis(4-tert-butylphenoxy)phosphoryl]-3-methylbutyl]-L-prolinamide
-
-
N-acetyl-L-tryptophan amide
molecular dynamics simulations are performed with subtilisin in the presence and in the absence of an inhibitor both in hexane and in water. The inhibitor induces an open conformation of the S1 pocket that is maintained after the removal of the ligand in anhydrous, but not in aqueous, simulations. The analysis of fluctuations suggest that this behavior is caused by the decreased flexibility exhibited by subtilisin in hexane
N-benzyloxycarbonyl-Ala-Pro-Phe-chloromethyl ketone
-
synthetic inhibitor
N-ethylmaleimide
at 37°C and pH of 7.5, 0.1 mM inhibits prosubtilisin JB1 by 15%
N1-18[ISP]
synthetic peptide corresponding to the N-terminal extension behaves as a mixed noncompetitive inhibitor of active ISP
pepstatin
-
almost completely inhibits acid proteolytic activity, does not inhibit alkaline proteolytic activity of purified Arp
pepstatin A
at 37°C and pH of 7.5, 0.1 mM inhibits prosubtilisin JB1 by 48%
propeptide
-
inhibition in a concentration-dependent manner. Wild-type propeptide is more potent than G56W-, G56S- and G56E-propeptide
-
tert-butyl (2S)-2-([1-[bis(2-methylphenoxy)phosphoryl]-3-methylbutyl]carbamoyl)pyrrolidine-1-carboxylate
-
-
tert-butyl (2S)-2-([1-[bis(3,4,5-trimethylphenoxy)phosphoryl]-3-methylbutyl]carbamoyl)pyrrolidine-1-carboxylate
-
-
tert-butyl (2S)-2-([1-[bis(3,4-dimethylphenoxy)phosphoryl]-3-methylbutyl]carbamoyl)pyrrolidine-1-carboxylate
-
-
tert-butyl (2S)-2-([1-[bis(4-methoxyphenoxy)phosphoryl]-3-methylbutyl]carbamoyl)pyrrolidine-1-carboxylate
-
-
tert-butyl (2S)-2-([1-[bis(4-methylphenoxy)phosphoryl]-3-methylbutyl]carbamoyl)pyrrolidine-1-carboxylate
-
-
tert-butyl (2S)-2-[(1-[bis[4-(sulfanylmethyl)phenoxy]phosphoryl]-2-phenylethyl)carbamoyl]pyrrolidine-1-carboxylate
-
-
tert-butyl (2S)-2-[[1-(diphenoxyphosphoryl)-2-phenylethyl]carbamoyl]pyrrolidine-1-carboxylate
-
-
tomato inhibitor-II
-
TI-II, enzyme binding structure, the interdomain interface in TI-II consists of a small cluster of highly conserved hydrophobic residues Ile14, Pro16, Tyr98, Phe100 and Phe106 from domain I and Tyr34, Pro54 and Lys55 from domain II. Although this interface is quite small (buried surface area of 487A), it forms a stable packing arrangement between the two domains. Each reactive site loop in TI-II interacts with a separate molecule of subtilisin in the canonical manner observed in other proteinase–inhibitor complexes. The domains of TI-II appear to bind the proteinase independently of each other
-
Triton X-100
at 37°C and pH of 7.5, 0.01% reduces prosubtilisin JB1 relative activity to 92% and 0.05% reduces prosubtilisin JB1 relative activity to 69%
Tween 20
at 37°C and pH of 7.5, 0.01% reduces prosubtilisin JB1 relative activity to 53% and 0.05% reduces prosubtilisin JB1 relative activity to 70%
Urea
high concentrations (6 M) of urea significantly decrease the enzyme activity by over 60%, most likely due to disrupting the enzyme structural integrity
1,4-dioxane
-
104fold decrease in the enzyme’s catalytic activity for the hydrolysis reaction of vinyl butyrate with D2O and a 50% decrease in enzyme structural dynamics. Attaching increasing amounts of PEG to the enzyme reverses some of the activity loss. Poly(ethylene)-glycolylation increases protein structural dynamics in 1,4-dioxane
1,4-dioxane
-
reduces the enzyme activity. Structural changes, flexibility, hydration, and changes of the enzyme ionization state are not responsible for the low storage stability. Possible depletion or rearrangement of water molecules around the active site, or small structural perturbations around the active site or movements of counter ions
antipain
at 37°C and pH of 7.5, 0.1 mM inhibits prosubtilisin JB1 by 75%
chymostatin
at 37°C and pH of 7.5, 0.1 mM inhibits prosubtilisin JB1 by 95%
Cu2+
at 37°C and pH of 7.5, with 1 mM results in an almost complete reduction of prosubtilisin JB1 activity, 5 mM reduces prosubtilisin JB1 relative activity to 19%
E-64
at 37°C and pH of 7.5, 0.1 mM inhibits prosubtilisin JB1 by 31%
EDTA
inactivates the enzyme at 0.01 mM, the enzyme protein becomes less structured and potentially monomeric. Removal of Ca2+ at sites close to the dimer interface and the S1 pocket are involved enzyme inhhibition by EDTA
EDTA
5 mM partially inhibits by 35%, in 100 mM Tris-HCl buffer, pH 7.0 at 15°C for 20 min
EDTA
at 37°C and pH of 7.5, 0.1 mM inhibits prosubtilisin JB1 by 52%
EDTA
-
subtilisin exhibits little activity at 80°C in the presence of 10 mM. Subtilisin-like serine protease is fully active even in the presence of 10 mM EDTA
Inhibitor from egg white
-
subtilisin Carlsberg, subtilisin BPN'
-
Inhibitor from seeds of Canavalia lineata
-
low MW protein of about 6500 MW
-
Inhibitor from seeds of Canavalia lineata
-
MW 22000, Kunitz type inhibitors
-
Inhibitor from Vigna unguiculata subsp. cylindrica
-
inactive with other enzymes
-
Inhibitor from Vigna unguiculata subsp. cylindrica
-
properties and kinetics of the inhibitor
-
Inhibitor of Amaranthus caudatus seeds
-
subtilisin Novo (BPN') from Bacillus amyloliquefaciens, subtilisin Carlsberg from Bacillus licheniformis
-
Inhibitor of Amaranthus caudatus seeds
-
subtilisin Novo (BPN') from Bacillus amyloliquefaciens, subtilisin Carlsberg from Bacillus licheniformis
-
nitrogen
-
together with carbon significantly reduces protease activity of KV01 at 16 and 36 h, and KV54 at 16 h
nitrogen
-
absence of C or N metabolite repression in subtilisins from KV42 isolate. Together with carbon significantly reduces protease activity of KV71 at 16 and 36 h, and KV22 at 16 h
phenylmethylsulfonyl fluoride
1 mM completely inhibits, in 100 mM Tris-HCl buffer, pH 7.0 at 15°C for 20 min
phenylmethylsulfonyl fluoride
at 37°C and pH of 7.5, 0.1 mM inhibits prosubtilisin JB1 by 79%
phenylmethylsulfonyl fluoride
-
inhibits alkaline proteolytic activity of purified Arp by 100%, does not inhibit acid proteolytic activity
PMSF
-
circular-dichroism and fluorescence studies of PMSF derivative of subtilisin indicate a closely similar structure to that of native subtilisin
Sodium dodecyl sulfate
-
activity of the enzyme is retarded by 2.3 and 244times in 1 mM and 40 mM sodium dodecyl sulfate respectively compared to that in buffer solution. No evidence of sandwich-like subtilisin–sodium dodecyl sulfate complex formation, thus the enzyme does not encroach into the hydrophobic surfactant core of sodium dodecyl sulfate micelle to form an elongated structure. Retains its structural integrity in sodium dodecyl sulfate solution. Micellar crowding in the vicinity of the enzyme
Sodium dodecyl sulfate
at 37°C and pH of 7.5, 0.01% reduces prosubtilisin JB1 relative activity to 52% and 0.05% reduces prosubtilisin JB1 relative activity to 29%
sucrose
-
together with carbon significantly reduces protease activity of KV01 at 16 and 36 h, and KV54 at 16 h
sucrose
-
absence of C or N metabolite repression in subtilisins from KV42 isolate. Together with carbon significantly reduces protease activity of KV71 at 16 and 36 h, and KV22 at 16 h
Zn2+
at 37°C and pH of 7.5, 1 mM reduces prosubtilisin JB1 relative activity to 30%, with 5 mM results in an almost complete reduction of prosubtilisin JB1 activity
additional information
-
not inhibitory: Tween-20, Tween-40, Tween-60, Tween-80 at 1%, sodium dodecylsulfate at 0.2%, for 1 h at 30°C. Not inhibitory: iodoacetate, ethylacetimide, phenylglyoxal, N-ethylmaleimide, N-bromosuccinimide
-
additional information
-
enzyme-specific, thermostable inhibitor from Physarum polycephalum with molecular mass of 32-33 kDa, 50% inhibition at 0.00014 mM
-
additional information
-
aldehyde and fluoromethyketone (FMK)-based inhibitors inhibit SAS-1 and -2 reversibly and only CMK-based inhibitors irreversibly inhibit the enzyme
-
additional information
-
molecular dynamics simulations, residue displacement correlations, and inhibitor design based on the enzyme inhibitor comlpex of TI-II and subtilisin, overview
-
additional information
-
metal chelating reagents; not: tosyl-leucine chloromethyl ketone; sulfhydryl reagents
-
additional information
-
strain DJ-4, no inhibition by EDTA or leupeptin
-
additional information
at 37°C and pH of 7.5, 0.1 mM leupeptin does not inhibit prosubtilisin JB1
-
additional information
enzyme destabilization in the presence of EDTA is due to chelation of a tightly bound calcium ion by EDTA, rather than destabilizing effects caused by EDTA on the structure
-
additional information
-
is not inhibited by 1,10-phenanthroline, EDTA, E64, bestatin, phosphoramidon and pepstatin A
-
additional information
-
alkaline proteolytic activity of purified Arp not inhibited by 3,4-dichloroisocumarin, tosyl lysine chloromethyl ketone, tosyl phenylalanine chloromethyl ketone, leupeptin, chymostatin, elastatinal, iodoacetamide, E-64, EDTA, 1,10-phenanthroline, aprotinin, antithrombin III, alpha1-antitrypsin and soybean trypsin inhibitor
-
additional information
the enzyme activity is not affected by various metal ions or non-specific protease inhibitors, e.g. bestatin, leupeptin, or pefabloc SC
-
additional information
-
enzymatic activity of mature subtilisin is inhibited by G56S-propeptide and wild-type propeptide in a concentration-dependent manner
-
additional information
-
subtilisin-like serine protease is fully resistant to treatment with 2-5% SDS, 4-8 M urea, 10% Tween-20 or 10% Triton X-100. In gel assay, subtilisin-like serine protease is fully denatured prior to SDS-PAGE by trichloroacetic acid treatment, followed by boiling for 5 min in the presence of SDS
-
additional information
no inhibition by EDTA and benzamidine
-
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
methyl-beta-cyclodextrin
-
activates and enhances the enantioselectivity of subtilisin C. mainly by prevention of structural perturbation during the initial lyophilization process. Minor contributions to increased activity likely stem from other factors, such as, reduction of possible mass transport limitations and changes in enzyme dynamics. Enhanced activity and enantioselectivity is not caused by formation of cyclodextrin–substrate or -product complexes
n-Propanol
-
subtilisin precipitated and rinsed with propanol yields higher transesterfication activity. About 10000times increase in initial rates in 1-butyl-3-methylimidazolium hexafluorophosphate over what is obtained with pH tuned lyophilized powders
OSP400
-
in the presence of 10 mM calcium the half-life of the enzyme at 60°C increases by 6.06fold. Among modifiers used OSP400 is most effective in stabilizing the enzyme. Thermally induced unfolding is delayed
-
OSP70
-
in the presence of 10 mM calcium the half-life of the enzyme at 60°C increases by 5.20fold. Thermally induced unfolding is delayed
-
poly(ethylene glycol)
-
poly(ethylene glycol)-modified subtilisin suspended in ionic liquids exhibits excellent catalytic performance while the native enzyme shows no activity
poly(ethyleneglycol)
-
presence of poly(ethyleneglycol) combined with trehalose during lyphilization does help in obtaining a more active enzyme preparation for catalysis in ionic liquids
polyglutaraldehyde
-
in the presence of 10 mM calcium the half-life of the enzyme at 60°C increases by 2.92fold. Thermally induced unfolding is delayed
[Emim][Tf2N]
-
poly(ethylene glycol)-modified subtilisin shows good stability in [Emim][Tf2N], and maintains 80% of its initial activity after 60 h. Preferred medium for enzymatic reaction. Enzyme activity is much higher than in conventional organic solvents. Excellent activity is associated with unique properties such as hydrophobicity and high polarity
-
additional information
-
prodomain mutant proR9 accelerates subtilisin folding. Subtilisin folding with the wild-type prodomain is slow. The wild-type prodomain binds 100times slower than proR9
-
additional information
-
conjugation of enzyme with comb-shaped poly-(ethylen glycol) and solubilization in ionic liquids without adding water. Enzyme exhibits higher transesterification activity in solution of [Eminm][Tf2N] than in toluene. No enzymic activity in DMSO, THF, or acetonitrile
-
additional information
-
lyophilized enzyme, sharp increase in enzyme activity in correlation with an increasing amount of fumed silica added to enzyme before freeze-drying. When freezing at -20°C instead of liquid nitrogen, even better activation is observed
-
additional information
-
improvement in the enzymatic activity for the poly(ethylene)-glycol-modified enzyme in the organic solvent that depends on the level of poly(ethylene)-glycolylation. PEG–SBc conjugates with 1.1, 1.9, and 3.2 average mol of poly(ethylene)-glycol per mol of protein reveal a 30fold, 57fold, and 99fold increase in enzymatic activity as a result of poly(ethylene)-glycolylation
-
additional information
-
no activity of poly(ethylene glycol)-modified subtilisin in methyl tert-butyl ether, tetrahydrofuran, dimethylsulfoxide, and acetonitrile
-
additional information
-
three phase partitioning reveals 13fold increase in the initial transesterfication rate
-
additional information
-
dynamics are also affected by the method of preparation of the enzyme: decreased flexibility is observed when the enzyme is chemically modified with poly ethylene glycol or colyophilized with crown ethers
-
additional information
-
sigma-H is required for full expression of the CSF-processing protease
-
additional information
-
glycosylation of subtilisin does not cause any significant tertiary structure changes
-
additional information
-
unlike subtilisin, subtilisin-like serine protease does not require propeptide for folding
-
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
8.3
benzyloxycarbonyl-Gly-4-nitroanilide
-
pH 7.5, 20°C, in presence and absence of DTT
-
90
N-Acetyl-Tyr methyl ester
-
subtilisin Novo, N-acetyl-Tyr ethyl ester, subtilisin Carlsberg, N-acetyl-Trp methyl ester, subtilisin Novo
0.000655
N-succinyl-L-Ala-L-Ala-L-Pro-L-Phe-4-nitroanilide
-
37°C, pH 10.5
0.36
succinyl-L-Ala-L-Ala-L-Phe-7-amido-4-methylcoumarin
-
mutant Y217L, pH 8.3, 25°C
0.45
succinyl-L-Ala-L-Ala-L-Pro-L-Phe-4-nitroanilide
-
mutant Y217L, pH 8.3, 25°C