Information on EC 1.14.13.154 - erythromycin 12-hydroxylase

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The expected taxonomic range for this enzyme is: Saccharopolyspora erythraea

EC NUMBER
COMMENTARY hide
1.14.13.154
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RECOMMENDED NAME
GeneOntology No.
erythromycin 12-hydroxylase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
erythromycin D + NADPH + H+ + O2 = erythromycin C + NADP+ + H2O
show the reaction diagram
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
erythromycin A biosynthesis
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megalomicin A biosynthesis
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Biosynthesis of 12-, 14- and 16-membered macrolides
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Biosynthesis of antibiotics
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SYSTEMATIC NAME
IUBMB Comments
erythromycin-D,NADPH:oxygen oxidoreductase (12-hydroxylating)
The enzyme is responsible for the C-12 hydroxylation of the macrolactone ring, one of the last steps in erythromycin biosynthesis. It shows 1200-1900-fold preference for erythromycin D over the alternative substrate erythromycin B [1].
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
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an Saccharopolyspora erythraea strain disrupted in eryK no longer produces erythromycin A but accumulates the B and D forms of the antibiotic
physiological function
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the enzyme is responsible for the C-12 hydroxylation of the macrolactone ring, one of the last steps in erythromycin biosynthesis
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
erythromycin B + NADPH + H+ + O2
erythromycin A + NADP+ + H2O
show the reaction diagram
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1200-1900-fold preference for erythromycin D over the alternative C-12 hydroxylase substrate erythromycin B
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?
erythromycin D + NADPH + H+ + O2
erythromycin C + NADP+ + H2O
show the reaction diagram
additional information
?
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in a Saccharopolyspora venezuelae pikC deletion mutant, EryK can catalyze the hydroxylation of YC-17 and narbomycin to generate methymycin/neomethymycin and pikromycin, respectively. EryK is flexible toward some alternative polyketides and can be useful for structural diversification of macrolides by post-polyketide synthase hydroxylation
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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
erythromycin D + NADPH + H+ + O2
erythromycin C + NADP+ + H2O
show the reaction diagram
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?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
cytochrome P-450
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a heme-thiolate protein (P-450)
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INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
clotrimazole
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induces a distortion of the internal helix I that affects the accessibility of the binding pocket by regulating the kink of the external helix G via a network of interactions that involves helix F
erythromycin D
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substrate inhibition
ketoconazole
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induces a distortion of the internal helix I that affects the accessibility of the binding pocket by regulating the kink of the external helix G via a network of interactions that involves helix F
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.008
erythromycin D
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pH 7.5, 30C
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.8
erythromycin D
Saccharopolyspora erythraea
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pH 7.5, 30C
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
225
erythromycin D
Saccharopolyspora erythraea
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pH 7.5, 30C
7926
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0166
erythromycin D
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pH 7.5, 30C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Saccharopolyspora erythraea (strain ATCC 11635 / DSM 40517 / JCM 4748 / NBRC 13426 / NCIMB 8594 / NRRL 2338)
Saccharopolyspora erythraea (strain ATCC 11635 / DSM 40517 / JCM 4748 / NBRC 13426 / NCIMB 8594 / NRRL 2338)
Saccharopolyspora erythraea (strain ATCC 11635 / DSM 40517 / JCM 4748 / NBRC 13426 / NCIMB 8594 / NRRL 2338)
Saccharopolyspora erythraea (strain ATCC 11635 / DSM 40517 / JCM 4748 / NBRC 13426 / NCIMB 8594 / NRRL 2338)
Saccharopolyspora erythraea (strain ATCC 11635 / DSM 40517 / JCM 4748 / NBRC 13426 / NCIMB 8594 / NRRL 2338)
Saccharopolyspora erythraea (strain ATCC 11635 / DSM 40517 / JCM 4748 / NBRC 13426 / NCIMB 8594 / NRRL 2338)
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
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x * 43759, calculated from sequence
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
crystal structures of EryK in complex with the azole inhibitors ketoconazole and clotrimazole
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different crystal forms are harvested from distinct crystallization conditions: two ligand-free forms, one substrate bound and two inhibitors-bound. All crystals belong either to the monoclinc P2(1)or to the orthorhombic P2(1)2(1)2(1) space groups, and exhibit diffraction limits ranging from 2.3 to 1.6 A
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vapor diffusion at 21C, three crystal forms of EryK are obtained in different crystallization conditions: with His tag (His6-EryK) in low salt conditions, without tag (EryK) in high salt, and in complex with its substrate erythromycin D (ErD-EryK)
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
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overproduction of EryK in Escherichia coli as insoluble inclusion bodies
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recombinant His-tagged EryK is overexpressed in BL21 STAR (DE3) Escherichia coli strain
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recombinant His-tagged EryK is overexpressed in the Escherichia coli BL21
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
synthesis
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systematically modulating the enzyme amounts of EryK and EryG by integrating additional eryK and eryG copies into the industrial strain Saccharopolyspora erythraea HL3168 E3 significantly enhances the process of biotransformation from erythromycin-D to erythromycin-A, nearly completely eliminates the by-products erythromycin-B and erythromycin-C, and efficiently improves erythromycin-A production and purity at the fermentation stage. In conjunction with other traditional and genetic ways to continuously evaluate the erythromycin-A production system