Information on EC 3.4.22.33 - Fruit bromelain

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The expected taxonomic range for this enzyme is: Bromeliaceae

EC NUMBER
COMMENTARY
3.4.22.33
-
RECOMMENDED NAME
GeneOntology No.
Fruit bromelain
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
Hydrolysis of proteins with broad specificity for peptide bonds. Bz-Phe-Val-Arg-/-NHMec is a good synthetic substrate, but there is no action on Z-Arg-Arg-NHMec (c.f. stem bromelain)
show the reaction diagram
-
-
-
-
REACTION TYPE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
hydrolysis of peptide bond
-
-
-
-
hydrolysis of peptide bond
-
-
hydrolysis of peptide bond
Q6Q2T4
preference for substrates possessing hydrophobic residues interacting with subsite S2, shows highest efficiency for Leu at P2
SYNONYMS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
Ananase
-
-
-
-
Bromelain
O23791
-
bromelain A
-
-
bromelain B
-
-
Bromelain, juice
-
-
-
-
Bromelase
-
-
-
-
Bromelin
-
-
-
-
Bromelin
-
-
EC 3.4.22.4
-
-
formerly
-
EC 3.4.22.5
-
-
formerly
-
Extranase
-
-
-
-
fastuosain
Q6Q2T4
-
Fruit bromelain
-
-
-
-
Fruit bromelain
-
-
Fruit bromelain FA2
-
-
-
-
Pinase
-
-
-
-
Traumanase
-
-
-
-
Juice bromelain
-
-
-
-
additional information
-
c.f. stem bromelain, enzyme belongs to the papain superfamily
additional information
-
another cysteine endopeptidase, with similar action on small molecule substrates, pinguinain (formerly EC 3.4.99.18) is obtained from the related plant, Bromelia pinguin, but pinguinain differs from fruit bromelain in being inhibited by chicken cystatin
CAS REGISTRY NUMBER
COMMENTARY
9001-00-7
-
ORGANISM
COMMENTARY
LITERATURE
SEQUENCE CODE
SEQUENCE DB
SOURCE
antigenic determinant
-
-
Manually annotated by BRENDA team
cultivar Smooth Cayenne
UniProt
Manually annotated by BRENDA team
mainly var. Cayenne
-
-
Manually annotated by BRENDA team
fragment
UniProt
Manually annotated by BRENDA team
another cysteine endopeptidase, with similar action on small molecule substrates, pinguinain (formerly EC 3.4.99.18), is obtained from the related plant, Bromelia pinguin, but pinguinain differs from fruit bromelain in being inhibited by chicken cystatin
-
-
Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                      
REACTION DIAGRAM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
+ H2O
?
show the reaction diagram
-
-
-
?
Angiotensin II + H2O
Hydrolyzed angiotensin II
show the reaction diagram
-
cleavage between Tyr4-Ile5
cleavage between Tyr4-Ile5
-
Angiotensin II + H2O
Hydrolyzed angiotensin II
show the reaction diagram
-
cleavage between Tyr4-Ile5
cleavage between Tyr4-Ile5
-
azocasein + H2O
?
show the reaction diagram
-
-
-
-
?
Benzoyl-Gly-Gly-Lys + H2O
?
show the reaction diagram
-
enzyme form FBA is much more active than FBB
-
-
-
benzoyl-L-arginine ethyl ester + H2O
?
show the reaction diagram
-
artificial substrate
-
-
?
benzoyl-Phe-Val-Arg-4-methylcoumarin 7-amide + H2O
benzoyl-Phe-Val-Arg + 7-amino-4-methylcoumarin
show the reaction diagram
-
convenient substrate for fruit bromelain, scarcely affected by stem bromelain
-
-
?
benzoyl-Phe-Val-Arg-p-nitroanilide + H2O
benzoyl-Phe-Val-Arg + p-nitroaniline
show the reaction diagram
-
-
-
-
?
Benzyloxycarbonyl-Ala-Leu + H2O
Benzyloxycarbonyl-Ala + Leu
show the reaction diagram
-
enzyme form FBA is much more active than FBB
-
-
-
benzyloxycarbonyl-Arg-Arg-p-nitroanilide + H2O
benzyloxycarbonyl-Arg-Arg + p-nitroaniline
show the reaction diagram
-
-
-
-
?
Benzyloxycarbonyl-Gly-Leu + H2O
Benzyloxycarbonyl-Gly + Leu
show the reaction diagram
-
enzyme form FBA is much more active than FBB
-
-
-
Benzyloxycarbonyl-Gly-Phe + H2O
Benzyloxycarbonyl-Gly + Phe
show the reaction diagram
-
-
-
-
-
Benzyloxycarbonyl-Gly-Tyr + H2O
Benzyloxycarbonyl-Gly + Tyr
show the reaction diagram
-
-
-
-
-
Benzyloxycarbonyl-Gly-Val + H2O
Benzyloxycarbonyl-Gly + Val
show the reaction diagram
-
enzyme form FBA is much more active than FBB
-
-
-
benzyloxyxarbonyl-Phe-Arg-7-amido-4-methylcoumarin + H2O
?
show the reaction diagram
-, Q6Q2T4
-
-
-
?
Bradykinin + H2O
Hydrolyzed bradykinin
show the reaction diagram
-
cleavage between Gly4-Phe5 and Phe5-Ser6
cleavage between Gly4-Phe5 and Phe5-Ser6
-
Bradykinin + H2O
Hydrolyzed bradykinin
show the reaction diagram
-
cleavage between Gly4-Phe5 and Phe5-Ser6
cleavage between Gly4-Phe5 and Phe5-Ser6
-
bromelain + H2O
?
show the reaction diagram
-
rapid self-digestion in aqueous solution, can be prevented by addition of alpha2-macroglobulin
-
?
Carbobenzoxy-Gly-Ala + H2O
Carbobenzoxy-Gly + Ala
show the reaction diagram
-
enzyme form FBA is much more active than FBB
-
-
-
casein + H2O
hydrolyzed casein
show the reaction diagram
-
-
-
-
-
casein + H2O
?
show the reaction diagram
-
-
-
?
casein + H2O
?
show the reaction diagram
-
-
-
-
?
casein + H2O
?
show the reaction diagram
O23791
-
-
-
?
casein + H2O
?
show the reaction diagram
-, Q6Q2T4
-
-
-
?
casein + H2O
?
show the reaction diagram
-
enzyme is assayed using casein as a substrate with Coomassie dye to track completion of hydrolysis of casein, casein is hydrolysed to completion leading to a brown coloured solution, assay method development as a Bradford dye-binding-bromelain-casein system as an assay for heavy metals, based on colour development and its inhibition by Hg2+ and Cu2+
-
-
?
epsilon-NH2-Cap-Ala-Cys(S-Bzl)-7-amido-4-methylcoumarin + H2O
?
show the reaction diagram
-, Q6Q2T4
-
-
-
?
epsilon-NH2-Cap-Cys(S-Bzl)-Leu-7-amido-4-methylcoumarin + H2O
?
show the reaction diagram
-, Q6Q2T4
-
-
-
?
epsilon-NH2-Cap-Leu-Cys(S-Bzl)-7-amido-4-methylcoumarin + H2O
?
show the reaction diagram
-, Q6Q2T4
-
-
-
?
epsilon-NH2-Cap-Leu-Gly-7-amido-4-methylcoumarin + H2O
?
show the reaction diagram
-, Q6Q2T4
-
-
-
?
epsilon-NH2-Cap-Phe-Arg-7-amido-4-methylcoumarin + H2O
?
show the reaction diagram
-, Q6Q2T4
-
-
-
?
epsilon-NH2-Cap-Pro-Cys(S-Bzl)-7-amido-4-methylcoumarin + H2O
?
show the reaction diagram
-, Q6Q2T4
-
-
-
?
Gelatin + H2O
?
show the reaction diagram
-
-
-
?
Gelatin + H2O
?
show the reaction diagram
-
-
-
-
?
Hemoglobin + H2O
Hydrolyzed hemoglobin
show the reaction diagram
-
-
-
-
-
N-alpha-benzoyl-DL-arginine-2-naphthylamide + H2O
N-alpha-benzoyl-DL-arginine + 2-naphthylamine
show the reaction diagram
-, Q6Q2T4
-
-
-
?
N-Benzoyl-L-Arg ethyl ester + H2O
N-Benzoyl-L-Arg + ethanol
show the reaction diagram
-
-
-
-
-
N-benzoyl-L-tyrosine ethyl ester + H2O
?
show the reaction diagram
-
artificial substrate
-
-
?
Nalpha-Benzoyl-Arg amide + H2O
Nalpha-Benzoyl-Arg + NH3
show the reaction diagram
-
-
-
-
-
Nalpha-Benzoyl-DL-Arg 4-nitroanilide + H2O
Nalpha-Benzoyl-DL-Arg + 4-nitroaniline
show the reaction diagram
-
-
-
-
-
Nalpha-benzoyl-L-arginine-4-nitroanilide + H2O
?
show the reaction diagram
-
artificial substrate
-
-
?
protein + H2O
peptides
show the reaction diagram
-
broad substrate specificity
-
?
Wheat gluten + H2O
Hydrolyzed wheat gluten
show the reaction diagram
-
-
-
-
-
Isolated soybean protein + H2O
Hydrolyzed soybean protein
show the reaction diagram
-
-
-
-
-
additional information
?
-
-
specificity overview
-
-
-
additional information
?
-
-
enzyme catalyzes synthesis of acylamino acid anilides
-
-
-
additional information
?
-
-
bromelain has a profound immunmodulatory effect, overview, bromelain affects blood coagulation, fibrinolysis, and platelet functions, mechanism, overview
-
?
additional information
?
-
-
enzyme is scarcely able to hydrolyze benzyloxycarbonyl-Arg-Arg-4-methylcoumarin 7-amide
-
-
-
additional information
?
-
-
bromelain is a cysteine protease containing a single thiol group in the active sites
-
-
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
protein + H2O
peptides
show the reaction diagram
-
broad substrate specificity
-
?
additional information
?
-
-
bromelain has a profound immunmodulatory effect, overview, bromelain affects blood coagulation, fibrinolysis, and platelet functions, mechanism, overview
-
?
METALS and IONS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
additional information
-
poor or no inhibition by ions K+, Ca2+, Mg2+, Mn2+, Co2+, Bo3+, Al3+, Cr2+, Ni2+, Sn2+, Ag+, Fe2+, Cd2+, Zn2+, W3+, Se2+, As3+, and Pb2+, overview
INHIBITORS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
Cu2+
Q6Q2T4
12% residual activity at 0.16 mM
Cu2+
-
inhibits casein degradation, complete inhibition
Cu2+
-
reduces activity
Cystatin
Q6Q2T4
-
-
Cystatin
O23791
an extended AE-rich N-terminal trunk in secreted pineapple cystatin enhances inhibition of fruit bromelain and is posttranslationally removed during ripening, The AE-rich N-terminal trunk is required to inhibit fruit bromelain (above 95%), whereas its removal decreases inhibition to 20%. Recombinant cystatin containing the complete AE-rich N-terminal trunk and recombinant medium pineapple cystatin effectively inhibit bromelain compared to recombinant core pineapple cystatin
-
Fe3+
-
reduces activity
-
Hg2+
Q6Q2T4
14% residual activity at 0.16 mM
Hg2+
-
inhibits casein degradation, complete inhibition
iodoacetic acid
Q6Q2T4
5% residual activity at 2 mM
Mercurials
-
cysteine restores activity
-
p-chloromercuribenzoate
Q6Q2T4
15% residual activity at 2 mM
PCMB
-
activity restored by cysteine
Phenylmethylsulfonylfluoride
Q6Q2T4
84% residual activity at 2 mM
Potato cysteine proteinase inhibitor
-
-
-
sulfhydryl reagents
-
-
Urea
Q6Q2T4
at 2.4 M urea the activity decreases continuously to 50% when using N-alpha-benzoyl-DL-arginine-2-naphthylamide as a substrate
Mercurials
-
cysteine restores activity; mercaptoethanol restores activity
-
additional information
-
scarcely inhibited by chicken chystatin
-
additional information
-
enzyme is inhibited by cystatin
-
additional information
Q6Q2T4
not inhibited by pepstatin
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
2-mercaptoethanol
Q6Q2T4
highest activity at 96 mM, at 37C and pH 7.0
dithiothreitol
Q6Q2T4
highest activity at 2.5 mM, at pH 7.0 and 37C
EDTA
Q6Q2T4
28% increase of activity at 2 mM
Urea
Q6Q2T4
activity decreases in the presence of 4 M urea and reaches the activity of the control again at 8 M when using casein as a substrate
KM VALUE [mM]
KM VALUE [mM] Maximum
SUBSTRATE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
43
-
N-benzoyl-L-Arg ethyl ester
-
-
2.1
-
Nalpha-Benzoyl-Arg amide
-
fruit bromelain C
4
-
Nalpha-Benzoyl-Arg amide
-
-
4
-
Nalpha-Benzoyl-Arg amide
-
fruit bromelain A and B
4
-
Nalpha-Benzoyl-Arg amide
-
-
TURNOVER NUMBER [1/s]
TURNOVER NUMBER MAXIMUM[1/s]
SUBSTRATE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
0.0094
-
Nalpha-Benzoyl-Arg amide
-
fruit bromelain C
0.027
-
Nalpha-Benzoyl-Arg amide
-
fruit bromelain B
0.033
-
Nalpha-Benzoyl-Arg amide
-
fruit bromelain A
pH OPTIMUM
pH MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
3.5
4
-
benzoyl-Glu-Tyr, benzoyl-Glu-Phe
3.5
-
-
benzoyl-Ala-Leu, benzoyl-Ala-Phe, enzyme form FBA
4
-
-
benzoyl-Ala-Leu, enzyme form FBA
6
-
-
assay at
6.5
-
-
benzoyl-Arg amide, enzyme form FBA
7
-
Q6Q2T4
-
7
-
-
assay at
8.3
-
-
casein
additional information
-
-
-
additional information
-
-
pI: 4.6
pH RANGE
pH RANGE MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
3.5
8.5
O23791
-
4
9
Q6Q2T4
-
TEMPERATURE OPTIMUM
TEMPERATURE OPTIMUM MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
30
-
-
assay at
37
-
-
assay at
45
-
-
assay at
55
-
Q6Q2T4
using casein as a subatrate
65
-
Q6Q2T4
using N-alpha-benzoyl-DL-arginine-2-naphthylamide as a substrate
TEMPERATURE RANGE
TEMPERATURE MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
50
60
-
most suitable hydrolysate temperature of casein acids for bromelain is 50-60C
pI VALUE
pI VALUE MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
4.6
4.7
-
isoelectric focusing
SOURCE TISSUE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
SOURCE
-
bromelain is a mixture of bromelain A and B, the proteolytic enzymes of pineapple fruit
Manually annotated by BRENDA team
-
stem bromelain possess better gelatin digestion units (GDU) activity over fruit bromelain
Manually annotated by BRENDA team
additional information
-
bromelain is a crude extract containing a complex mixture of several components, such as different thiol-endopeptidases, phosphatases, glucosidases, peroxidases, cellulases, glycoproteins and carbohydrates, as well as several proteinase inhibitors
Manually annotated by BRENDA team
MOLECULAR WEIGHT
MOLECULAR WEIGHT MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
18000
-
-
Ananas comosus, fruit bromelain A, gel filtration
19000
-
-
Ananas comosus, fruit bromelain C, gel filtration
25000
-
Q6Q2T4
SDS-PAGE
28000
-
-
Ananas comosus, fruit bromelain B, gel filtration
28000
-
O23791
SDS-PAGE
30000
-
-
SDS-PAGE
31000
-
-
Ananas comosus, gel electrophoresis
32500
-
-
Ananas comosus, gel filtration
34000
-
-
SDS-PAGE
SUBUNITS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
?
-
x * 23000, Ananas comosus, enzyme form FBA, SDS-PAGE, x * 27000, Ananas comosus, enzyme form FBB, SDS-PAGE
?
-
x * 23000, SDS-PAGE
monomer
-
1 * 31000, Ananas comosus, SDS-PAGE, 1 * 32500, Ananas comosus
monomer
-
-
monomer
-
1 * 25000
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
no glycoprotein
-
-
pH STABILITY
pH STABILITY MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
2
-
-
at a concentration of 250 mg/ml the enzyme is resistant to inactivation
TEMPERATURE STABILITY
TEMPERATURE STABILITY MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
60
-
-
inactivation increases if temperature exceeds 60C
GENERAL STABILITY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
incubation with trypsin (1 mg/ml, 1 h, 37C) results in 12% loss of activity (with benzyloxycarbonyl-Arg-Arg-p-nitroanilide as substrate) at bromelain concentration of 10 mg/ml , but has no effect on activity at bromelain concentrations of 100 mg/ml
-
proteolytic activity of solutions containing either 10 mg/ml or 100 mg/ml bromelain in water or 100 mg/ml NaHCO3 is not changed against benzyloxycarbonyl-Arg-Arg-p-nitroanilide and benzoyl-Phe-Val-Arg-p-nitroanilide substrate by four freeze/thaw cycles
-
Purification/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
2 forms: FBA and FBB; fractionation
-
bromelain from pineapple stem and skin are recovered by a PEG 4000/phosphate aqueous two-phase systems (ATPs) liquid-liquid extraction
-
fractionation; fruit bromelain A, B and C
-
high-speed counter-current chromatography coupled with the reverse micelle solvent system consisting of 0.10 g/ml cetyltrimethylammonium bromide (CTAB)/isooctane-hexylalcohol and 0.05 M sodium phosphate buffer is successfully applied to separate bromelain from pineapple fruit
-
reverse micellar extraction system with cetyltrimethylammonium bromide
O23791
using centrifugation, salt precipitation technique, dialysis, ion-exchange chromatography. Ion exchange chromatography using diethylaminoethyl cellulose (DEAE) anion exchangers maintain the structural integrity of purified bromelain and thereby the product exhibits better proteolytic activity than crude extract
-
CM-Sepharose column chromatography, SP-Sepharose column chromatography, and Sephadex G-50 gel filtration
Q6Q2T4
Cloned/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
expressed in Escherichia coli Top10 cells
Q6Q2T4
APPLICATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
medicine
-
bromelain can be used as a phytotherapy agent in several aspects, overview
pharmacology
-
use as additive to prevent lymphedema by reducing lymphocongestion, detritus, viscosity of the exsudate and stimulation of phagocytosis of associated leukocytes, use as additive for radiotherapy and surgery due to wound healing effect and reductive effect on inflammation and edema, use as additives for chemotherapy due to inhibitory effects on malignant growth of tumor cells