Information on EC 3.4.22.71 - sortase B

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The expected taxonomic range for this enzyme is: Bacilli

EC NUMBER
COMMENTARY hide
3.4.22.71
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RECOMMENDED NAME
GeneOntology No.
sortase B
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
The enzyme catalyses a cell wall sorting reaction in which a surface protein with a sorting signal containing a NXTN motif is cleaved. The resulting threonine carboxyl end of the protein is covalently attached to a pentaglycine cross-bridge of peptidoglycan.
show the reaction diagram
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of peptide bond
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CAS REGISTRY NUMBER
COMMENTARY hide
9033-39-0
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain Newman
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
Q2FHU6
the enzyme is a member of the sortase cysteine transpeptidase family. Members of this enzyme superfamily are widely distributed in Gram-positive bacteria that frequently utilize multiple sortases to elaborate their peptidoglycan, and the family members have a conserved active site His-Cys-Arg triad that joins a sorting signal located at the C-terminus of their protein substrate to an amino nucleophile located on the cell surface
physiological function
additional information
Q2FHU6
analysis of the substrate binding structure of the enzyme using SrtB-NPQT complexes, by computational modeling, molecular dynamics simulations, and targeted amino acid mutagenesis revealing that the backbone amide of Glu224 and the side chain of Arg233 form an oxyanion hole in sortase B that stabilizes high energy tetrahedral catalytic intermediates. A highly conserved threonine residue within the bound sorting signal substrate facilitates construction of the oxyanion hole by stabilizing the position of the active site arginine residue via hydrogen bonding
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
Abz-KDFEVPTGVAM-Dap(Dnp)-NH2 + H2O
?
show the reaction diagram
IsdC + H2O
?
show the reaction diagram
Lmo2185 + H2O
?
show the reaction diagram
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?
Lmo2186 + H2O
?
show the reaction diagram
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NPKSS is a sorting motif of Lmo2186. Recognition of NPKSS by SrtB, even when placed in the context of the heterologous sorting signal of Lmo2185. Proline at position 2, and not lysine at position 3, is essential for the recognition of NPKSS by SrtB
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?
SvpA + H2O
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show the reaction diagram
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anchoring of SvpA to the bacterial cell wall is specifically mediated by SrzB
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?
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
IsdC + H2O
?
show the reaction diagram
SvpA + H2O
?
show the reaction diagram
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anchoring of SvpA to the bacterial cell wall is specifically mediated by SrzB
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?
additional information
?
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Zn2+
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the asymmetric unit contains 19 bound Zn2+ ions
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
(Z)-3-(2,5-dimethoxyphenyl)-2-(4-methoxyphenyl) acrylonitrile
berberine chloride
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potential of this inhibitor for the treatment of Staphylococcus aureus infections
beta-sitosterol-3-O-glucopyranoside
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potential of this inhibitor for the treatment of Staphylococcus aureus infections
galangin
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IC50 for recombinant SrtB(DELTA30): 0.03837 mM, no antibacterial activity against Staphylococcus aureus
galangin-3-methyl ether
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IC50 for recombinant SrtB(DELTA30): 0.1136 mM, no antibacterial activity against Staphylococcus aureus
isorhamnetin
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IC50 for recombinant SrtB(DELTA30): 0.04335 mM, no antibacterial activity against Staphylococcus aureus
kaempferol
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IC50 for recombinant SrtB(DELTA30): 0.02455 mM, no antibacterial activity against Staphylococcus aureus
morin
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IC50 for recombinant SrtB(DELTA30): 0.00854 mM, no antibacterial activity against Staphylococcus aureus
myricetin
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IC50 for recombinant SrtB(DELTA30): 0.03689 mM, no antibacterial activity against Staphylococcus aureus
psammaplin A1
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potential of this inhibitor for the treatment of Staphylococcus aureus infections
quercetin
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IC50 for recombinant SrtB(DELTA30): 0.03328 mM, no antibacterial activity against Staphylococcus aureus
quercetin-3,3'-dimethyl ether
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IC50 for recombinant SrtB(DELTA30): 0.0603 mM, no antibacterial activity against Staphylococcus aureus
additional information
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aryl (beta-amino)ethyl ketones inhibit sortase enzymes. Inhibition of sortases occurs through an irreversible, covalent modification of their active site cysteine. Sortases specifically activate this class of molecules via beta-elimination, generating a reactive olefin intermediate that covalently modifies the cysteine thiol
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IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.03837
galangin
Staphylococcus aureus
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IC50 for recombinant SrtB(DELTA30): 0.03837 mM, no antibacterial activity against Staphylococcus aureus
0.1136
galangin-3-methyl ether
Staphylococcus aureus
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IC50 for recombinant SrtB(DELTA30): 0.1136 mM, no antibacterial activity against Staphylococcus aureus
0.04335
isorhamnetin
Staphylococcus aureus
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IC50 for recombinant SrtB(DELTA30): 0.04335 mM, no antibacterial activity against Staphylococcus aureus
0.02455
kaempferol
Staphylococcus aureus
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IC50 for recombinant SrtB(DELTA30): 0.02455 mM, no antibacterial activity against Staphylococcus aureus
0.00854
morin
Staphylococcus aureus
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IC50 for recombinant SrtB(DELTA30): 0.00854 mM, no antibacterial activity against Staphylococcus aureus
0.03689
myricetin
Staphylococcus aureus
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IC50 for recombinant SrtB(DELTA30): 0.03689 mM, no antibacterial activity against Staphylococcus aureus
0.03328
quercetin
Staphylococcus aureus
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IC50 for recombinant SrtB(DELTA30): 0.03328 mM, no antibacterial activity against Staphylococcus aureus
0.0603
quercetin-3,3'-dimethyl ether
Staphylococcus aureus
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IC50 for recombinant SrtB(DELTA30): 0.0603 mM, no antibacterial activity against Staphylococcus aureus
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7
Q2FHU6
assay at
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
37
Q2FHU6
assay at
PDB
SCOP
CATH
ORGANISM
UNIPROT
Peptoclostridium difficile (strain 630)
Staphylococcus aureus (strain USA300)
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
1.6 A resolution. Space group P2(1) with cell dimension of a = 40.47 A, b = 64.6 A, c = 42.96 A, alpha = 105.77°, beta = 105.77°, gamma = 90°
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SrtB in complex with aryl (beta-amino)ethyl ketone inhibitors. Analysis of the three-dimensional structure ofBacillusanthracissortaseBwithandwithoutinhibitorprovidesinsights into the mechanism of inhibition
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2.0 A resolution. Space group P2(1)2(1)2(1) with cell dimension of a = 71.208 A, b = 104.367 A, c = 58.087 A, alpha = beta = gamma = 90°
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hanging-drop vapor diffusion method. Crystal structure of SrtB-DELTA-N30 in complex with two active site inhibitors E64 and MTSET, and with the cell wall substrate analog tripleglycine
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sortase B enzyme in a covalent complex with an analogue of its NPQTN sorting signal substrate, hanging drop vapour diffusion method, micing of 0.150 mMSrtB-NPQT in 10 mM Tris-HCl, pH 7.0, 20 mM NaCl, with reservoir solution containing 2.8 M ammonium sulfate, 70 mM sodium citrate, pH 5.0, X-ray diffraction structure determination and analysis at 2.49 A resolution, molecular replacement method
Q2FHU6
hanging or sitting drop vapor diffusion method, using 10% (w/v) polyethylene glycol 3350, 0.2 M Zn(OAc)2, 0.1 M NaOAc, pH 6.2
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Ni-charged HiTrap column chromatography and Superdex 200 gel filtration
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recombinant wild-type and mutant enzymes from Escherichia coli strain Rosetta (DE3) pLysS by affinity chromatography
Q2FHU6
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli BL21 (DE3) pRP cells
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expression in Escherichia coli
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recombinant expression of wild-type and mutant enzymes in Escherichia coli strain Rosetta (DE3) pLysS
Q2FHU6
SrtADELTA24 is expressed in Escherichia coli
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C223A
Q2FHU6
site-directed mutagenesis, inactive mutant
D225A
Q2FHU6
site-directed mutagenesis, the mutant exhibits nonspecific proteolytic activity
H130A
Q2FHU6
site-directed mutagenesis, inactive mutant
R233A
Q2FHU6
site-directed mutagenesis, inactive mutant
additional information
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marked changes in the specificity profile of SrtA are obtained by replacing the beta6/beta7 loop in SrtA with the corresponding domain from SrtB
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
medicine
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potential to be used as inhibitors for the treatment of Staphylococcus aureus infections: (Z)-3-(2,5-dimethoxyphenyl)-2-(4-methoxyphenyl) acrylonitrile, beta-sitosterol-3-O-glucopyranoside, berberine chloride and psammaplin A1