Information on EC 2.3.1.164 - isopenicillin-N N-acyltransferase

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The expected taxonomic range for this enzyme is: Trichocomaceae

EC NUMBER
COMMENTARY hide
2.3.1.164
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RECOMMENDED NAME
GeneOntology No.
isopenicillin-N N-acyltransferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
phenylacetyl-CoA + isopenicillin N + H2O = CoA + penicillin G + L-2-aminohexanedioate
show the reaction diagram
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-
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Acylation
hydrolysis
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Biosynthesis of antibiotics
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Metabolic pathways
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Penicillin and cephalosporin biosynthesis
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penicillin K biosynthesis
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SYSTEMATIC NAME
IUBMB Comments
acyl-CoA:isopenicillin N N-acyltransferase
Proceeds by a two stage mechanism via 6-aminopenicillanic acid. Different from EC 3.5.1.11, penicillin amidase.
CAS REGISTRY NUMBER
COMMENTARY hide
54576-90-8
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
gene penDE
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Manually annotated by BRENDA team
Wis 54-1255
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Manually annotated by BRENDA team
high yielding strain
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
metabolism
physiological function
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
phenoxyacetyl-CoA + isopenicillin N + H2O
CoA + penicillin V + L-2-aminohexanedioate
show the reaction diagram
phenylacetyl-CoA + 6-aminopenicillanic acid + H2O
CoA + benzylpenicillin
show the reaction diagram
phenylacetyl-CoA + isopenicillin N + H2O
CoA + benzylpenicillin + L-2-aminohexanedioate
show the reaction diagram
phenylacetyl-CoA + isopenicillin N + H2O
CoA + penicillin G + L-2-aminohexanedioate
show the reaction diagram
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
phenylacetyl-CoA + isopenicillin N + H2O
CoA + benzylpenicillin + L-2-aminohexanedioate
show the reaction diagram
-
-
-
?
phenylacetyl-CoA + isopenicillin N + H2O
CoA + penicillin G + L-2-aminohexanedioate
show the reaction diagram
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
8
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activity assay
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
26
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activity assay
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
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IAT encoded by the aatB gene, lacking a peroxisomal targeting signal PTS1
Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
11000
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processed protein, subunit alpha, determined by SDS-PAGE and Western blot analysis
28000
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SDS-PAGE
29000
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processed protein, subunit beta, determined by SDS-PAGE and Western blot analysis
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
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recombinant expression in Hansenula polymorpha give a 40 kDa pre-protein that is processed into an 11 and 29 kDa subunit, SDS-PAGE
heterodimer
additional information
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
proteolytic modification
additional information
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ptoIAT overexpressed in Escherichia coli is not self-processed even when treated with Aspergillus nidulans crude extract
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
hanging-drop vapour-diffusion method, X-ray diffraction, monoclinic space group C2, unit-cell parameteres: a = 231.36 A, b = 68.27 A, c = 151.31 A, beta = 129.6° at -173°C
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cell extracts of Penicillium chrysogenum are prepared, Escherichia coli soluble extracts are used for in vitro IAT activity measurement
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recombinant wild-type enzyme with a C-terminal His-tag is purified in a single step using Ni2+ affinity chelate chromatography. A combination of 5 mM DTT + 250 mM NaCl + 5 mM EDTA is effective in preventing aggregation of the enzyme
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli JM109
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expression in Escherichia coli
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gene penDE, lack of correlation between the protein and transcript levels of the IAT enzyme, quantitative real-time PCR expression analysis and gene copy number determination, overexpression of penDE in Penicillium chrysogenum strains DS47274, DS47273, and DS17690, introduction of penDE gene without introns into these strains in a random manner, via the nonhomologous end-joining pathway. Cooverexpression of penDE with gene phl, encoding the phenylacetic acid CoA ligase leading to production of phenoxyacetyl-CoA and penicillin V
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overexpression in Escherichia coli, not self-processed even when treated with Aspergillus nidulans crude extract
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pPenIAT and pPenIATC103S are used to express the penDE gene in Penicillium chrysogenum, pPBCalphabeta, pDEC103S and palphabeta-DEC103S are used for expression in Escherichia coli
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the intronless gene is expressed in Escherichia coli and self-processed efficiently into its alpha and beta subunits
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
transcription factor AnBH1 acting as a repressor of aatA and aatB expression
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transcription factor AnCF acting as an activator of aatA and aatB expression
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C103S
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unprocessed inactive variant of isopenicillin N acyltransferase
C103S or A or W
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C103 is specifically required for recombinant proenzyme cleavage
S227A
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produces uncleaved proenzyme lacking enzyme activity
S309A
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recombinant proenzyme containing S309A is cleaved, however enzyme activity is not observed
T105V
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results in a significant inhibition of proenzyme cleavage
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
biotechnology
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Penicillium chrysogenum npe6 lacking isopenicillin N acyltransferase activity is an excellent host for production of different beta-lactam antibiotics