PDBsum entry 1e5h

Oxidoreductase

PDB id: 1e5h

Contents

Protein chain

271 a.a. *

Ligands

SIN

CO2

Metals

FE2

Waters ×119

* Residue conservation analysis

PDB id:

1e5h

Name:

Oxidoreductase

Title:

Delta-r307a deacetoxycephalosporin c synthase complexed with succinate and carbon dioxide

Structure:

Deacetoxycephalosporin c synthase. Chain: a. Synonym: ring expanding enzyme, ring expandase. Engineered: yes. Mutation: yes

Source:

Streptomyces clavuligerus. Organism_taxid: 1901. Gene: cefe. Expressed in: escherichia coli. Expression_system_taxid: 469008. Other_details: recombinant e.Coli

Biol. unit:

Trimer (from PDB file)

Resolution:

1.96Å R-factor: 0.220 R-free: 0.252

Authors:

H.J.Lee,M.D.Lloyd,K.Harlos,I.J.Clifton,J.E.Baldwin,C.J.Schofield

Key ref:

H.J.Lee et al. (2001). Kinetic and crystallographic studies on deacetoxycephalosporin C synthase (DAOCS). J Mol Biol, 308, 937-948. PubMed id: 11352583 DOI: 10.1006/jmbi.2001.4649

Date:

26-Jul-00 Release date: 26-Jul-01

Protein chain

P18548  (CEFE_STRCL) -  Deacetoxycephalosporin C synthase from Streptomyces clavuligerus

Seq: 311 a.a.

Struc: 271 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.1.14.20.1 - deacetoxycephalosporin-C synthase.
• Pathway: Penicillin N and Deacetoxycephalosporin C Biosynthesis
• Reaction: penicillin N + 2-oxoglutarate + O2 = deacetoxycephalosporin C + succinate + CO2 + H2O

penicillin N + 2-oxoglutarate + O2 = deacetoxycephalosporin C (Bound ligand (Het Group name = CO2) corresponds exactly) + succinate + CO2 + H2O (Bound ligand (Het Group name = SIN) corresponds exactly)

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

DOI no: 10.1006/jmbi.2001.4649

J Mol Biol 308:937-948 (2001)

PubMed id: 11352583

Kinetic and crystallographic studies on deacetoxycephalosporin C synthase (DAOCS).

H.J.Lee, M.D.Lloyd, K.Harlos, I.J.Clifton, J.E.Baldwin, C.J.Schofield.

ABSTRACT

Deacetoxycephalosporin C synthase (DAOCS) is an iron(II) and 2-oxoglutarate-dependent oxygenase that catalyzes the conversion of penicillin N to deacetoxycephalosporin C, the committed step in the biosynthesis of cephalosporin antibiotics. The crystal structure of DAOCS revealed that the C terminus of one molecule is inserted into the active site of its neighbor in a cyclical fashion within a trimeric unit. This arrangement has hindered the generation of crystalline enzyme-substrate complexes. Therefore, we constructed a series of DAOCS mutants with modified C termini. Oxidation of 2-oxoglutarate was significantly uncoupled from oxidation of the penicillin substrate in certain truncated mutants. The extent of uncoupling varied with the number of residues deleted and the penicillin substrate used. Crystal structures were determined for the DeltaR306 mutant complexed with iron(II) and 2-oxoglutarate (to 2.10 A) and the DeltaR306A mutant complexed with iron(II), succinate and unhydrated carbon dioxide (to 1.96 A). The latter may mimic a product complex, and supports proposals for a metal-bound CO(2) intermediate during catalysis.

Selected figure(s)

Figure 1.
Figure 1. Structure of (a) iron(II) and 2-oxoglutarate complex of wild-type enzyme [Valegard et al 1998]; (b) iron(II) and 2-oxoglutarate complex of the DR306 mutant; and (c) iron(II), succinate and CO[2] complex of the DR307A mutant. The jelly-roll motif is shown in red, C termini are shown in blue, and the C terminus from symmetry related molecules in green.

Figure 3.
Figure 3. A view of the structure of the DR307A mutant iron(II), succinate and CO[2] complex. (a) A similar view to that in Figure 2, with the above residues omitted for clarity. The electron density map is contoured to 1.29s; (b) close up view of the CO[2]-binding site with succinate and the side-chains of Val262 and Phe264 enclosing one face. The remaining face of the molecule is bordered by residues in the C terminus, the conformation of which may be significantly different in solution (see the text for a discussion). The electron density map is contoured to 1.69s.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2001, 308, 937-948) copyright 2001.

Figures were selected by an automated process.