MetaCyc Enzyme: deacetoxycephalosporin C synthase

Gene: cefE Accession Number: G-10200 (MetaCyc)

Species: Streptomyces clavuligerus

Penicillin N is a precursor of the β-lactam antibiotics cephamycin C and cephalosporin C. Cephamycin C and other cephamycins (7-methoxycephalosporins) are biosynthesized by bacteria such as Streptomyces clavuligerus. Cephalosporin C and other cephalosporins are biosynthesized by fungi such as Acremonium chrysogenum (previously known as Cephalosporium acremonium). See superpathway of penicillin, cephalosporin and cephamycin biosynthesis.

In S. clavuligerus two separate enzymes, deacetoxycephalosporin C synthase and deacetoxycephalosporin C hydroxylase, are used to catalyze two sequential reactions that lead to the biosynthesis of cephamycin C. These two enzymes are encoded by the cef E and cefF genes, respectively. In contrast, A. chrysogenum uses a bifunctional enzyme deacetoxycephalosporin C synthase / deacetoxycephalosporin C hydroxylase to catalyze these two reactions that lead to the biosynthesis of cephalosporin C in this organism. This bifunctional enzyme is encoded by the cefEF gene. Reviewed in [Brakhage98] and [Thykaer03]

Both native and recombinant S. clavuligerus enzymes were found to have similar biochemical properties. Data given here are for the recombinant enzyme [Dotzlaf89].

Gene Citations: [Kovacevic89, Kovacevic90]

Molecular Weight of Polypeptide: 34.556 kD (from nucleotide sequence)

Unification Links: Protein Model Portal:P18548 , SMR:P18548 , UniProt:P18548

Relationship Links: Entrez-Nucleotide:RELATED-TO:M32324 , InterPro:IN-FAMILY:IPR002057 , InterPro:IN-FAMILY:IPR005123 , InterPro:IN-FAMILY:IPR026992 , InterPro:IN-FAMILY:IPR027443 , PDB:Structure:1DCS , PDB:Structure:1E5H , PDB:Structure:1E5I , PDB:Structure:1HJF , PDB:Structure:1HJG , PDB:Structure:1RXF , PDB:Structure:1RXG , PDB:Structure:1UNB , PDB:Structure:1UO9 , PDB:Structure:1UOB , PDB:Structure:1UOF , PDB:Structure:1UOG , PDB:Structure:1W28 , PDB:Structure:1W2A , PDB:Structure:1W2N , PDB:Structure:1W2O , PDB:Structure:2JB8 , Pfam:IN-FAMILY:PF03171 , Pfam:IN-FAMILY:PF14226 , Prosite:IN-FAMILY:PS00186 , Prosite:IN-FAMILY:PS51471

Gene-Reaction Schematic: ?

Gene-Reaction Schematic

Created 20-Sep-2007 by Fulcher CA , SRI International

Enzymatic reaction of: deacetoxycephalosporin C synthase

EC Number:

2-oxoglutarate + penicillin N + oxygen <=> CO2 + succinate + deacetoxycephalosporin C + H2O

The reaction direction shown, that is, A + B ↔ C + D versus C + D ↔ A + B, is in accordance with the direction in which it was curated.

The reaction is physiologically favored in the direction shown.

In Pathways: superpathway of penicillin, cephalosporin and cephamycin biosynthesis , deacetylcephalosporin C biosynthesis

Deacetoxycephalosporin C synthase (expandase) catalyzes the conversion of penicillin N to deacetoxycephalosporin C. This reaction involves ring expansion. The five-membered penam thiazolidine ring is oxidatively opened and after closure, the six-membered dihydrothiazine ring is formed. This ring is characteristic of ceph-3-ems. The enzyme is an α-ketoglutarate (2-oxoglutarate) linked dioxygenase. Reviewed in [Brakhage98] and [Thykaer03]

Neither the native, nor the recombinant enzyme was stimulated by ATP, in contrast to the Acremonium chrysogenum enzyme. The recombinant enzyme also catalyzed a novel hydroxylation of 3-exomethylenecephalosporin-C to deacetylcephalosporin C similar to the Acremonium chrysogenum enzyme, but did not catalyze the hydroxylation of deacetoxycephalosporin C. 3-exomethylenecephalosporin-C competitively inhibited the enzyme with respect to penicillin N [Dotzlaf89].

Cofactors or Prosthetic Groups: 2-oxoglutarate [Dotzlaf89], Fe2+ [Dotzlaf89], oxygen [Dotzlaf89]

Activators (Unknown Mechanism): L-dithiothreitol [Dotzlaf89] , L-ascorbate [Dotzlaf89]

Inhibitors (Competitive): 3-exomethylenecephalosporin-C [Dotzlaf89]

Inhibitors (Unknown Mechanism): penicillin G [Dotzlaf89] , penicillin V [Dotzlaf89] , 5,5'-dithio-bis-2-nitrobenzoate [Dotzlaf89] , N-ethylmaleimide [Dotzlaf89] , o-phenanthroline [Dotzlaf89] , p-hydroxymercuribenzoate [Dotzlaf89] , EDTA [Dotzlaf89]

Kinetic Parameters:

Km (μM)
penicillin N

T(opt): 36 °C [Dotzlaf89]

pH(opt): 7.0 [Dotzlaf89]


Brakhage98: Brakhage AA (1998). "Molecular regulation of beta-lactam biosynthesis in filamentous fungi." Microbiol Mol Biol Rev 62(3);547-85. PMID: 9729600

Dotzlaf89: Dotzlaf JE, Yeh WK (1989). "Purification and properties of deacetoxycephalosporin C synthase from recombinant Escherichia coli and its comparison with the native enzyme purified from Streptomyces clavuligerus." J Biol Chem 264(17);10219-27. PMID: 2656705

Kovacevic89: Kovacevic S, Weigel BJ, Tobin MB, Ingolia TD, Miller JR (1989). "Cloning, characterization, and expression in Escherichia coli of the Streptomyces clavuligerus gene encoding deacetoxycephalosporin C synthetase." J Bacteriol 171(2);754-60. PMID: 2644235

Kovacevic90: Kovacevic S, Tobin MB, Miller JR (1990). "The beta-lactam biosynthesis genes for isopenicillin N epimerase and deacetoxycephalosporin C synthetase are expressed from a single transcript in Streptomyces clavuligerus." J Bacteriol 172(7);3952-8. PMID: 1694525

Thykaer03: Thykaer J, Nielsen J (2003). "Metabolic engineering of beta-lactam production." Metab Eng 5(1);56-69. PMID: 12749845

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Please cite the following article in publications resulting from the use of MetaCyc: Caspi et al, Nucleic Acids Research 42:D459-D471 2014
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