2BJS

Isopenicillin N synthase C-terminal truncation mutant

Summary for 2BJS

• Entry DOI: 10.2210/pdb2bjs/pdb
• Related: 1BK0 1BLZ 1HB1 1HB2 1HB3 1HB4 1IPS 1OBN 1OC1 1ODM 1ODN 1QIQ 1QJE 1QJF 1UZW 1W03 1W04 1W05 1W06 1W3V 1W3X 2BU9
• Descriptor: ISOPENICILLIN N SYNTHETASE, FE (III) ION, L-D-(A-AMINOADIPOYL)-L-CYSTEINYL-D-VALINE, ... (6 entities in total)
• Functional Keywords: oxidoreductase, antibiotic biosynthesis, b-lactam antibiotic, iron, oxygenase, penicillin biosynthesis, vitamin c
• Biological source: Emericella nidulans (strain FGSC A4 / ATCC 38163 / CBS 112.46 / NRRL 194 / M139) (Aspergillus nidulans)
• Total number of polymer chains: 1
• Total formula weight: 37847.04

Authors

McNeill, L.A.,Sami, M.,Clifton, I.J.,Burzlaff, N.I. (deposition date: 2005-02-07, release date: 2006-03-09, Last modification date: 2024-05-08)

Primary citation

McNeill, L.A.,Brown, T.J.N.,Sami, M.,Clifton, I.J.,Burzlaff, N.I.,Claridge, T.D.W.,Adlington, R.M.,Baldwin, J.E.,Rutledge, P.J.,Schofield, C.J.
Terminally Truncated Isopenicillin N Synthase Generates a Dithioester Product: Evidence for a Thioaldehyde Intermediate during Catalysis and a New Mode of Reaction for Non-Heme Iron Oxidases.
Chemistry, 23:12815-12824, 2017

PubMed Abstract: Isopenicillin N synthase (IPNS) catalyses the four-electron oxidation of a tripeptide, l-δ-(α-aminoadipoyl)-l-cysteinyl-d-valine (ACV), to give isopenicillin N (IPN), the first-formed β-lactam in penicillin and cephalosporin biosynthesis. IPNS catalysis is dependent upon an iron(II) cofactor and oxygen as a co-substrate. In the absence of substrate, the carbonyl oxygen of the side-chain amide of the penultimate residue, Gln330, co-ordinates to the active-site metal iron. Substrate binding ablates the interaction between Gln330 and the metal, triggering rearrangement of seven C-terminal residues, which move to take up a conformation that extends the final α-helix and encloses ACV in the active site. Mutagenesis studies are reported, which probe the role of the C-terminal and other aspects of the substrate binding pocket in IPNS. The hydrophobic nature of amino acid side-chains around the ACV binding pocket is important in catalysis. Deletion of seven C-terminal residues exposes the active site and leads to formation of a new type of thiol oxidation product. The isolated product is shown by LC-MS and NMR analyses to be the ene-thiol tautomer of a dithioester, made up from two molecules of ACV linked between the thiol sulfur of one tripeptide and the oxidised cysteinyl β-carbon of the other. A mechanism for its formation is proposed, supported by an X-ray crystal structure, which shows the substrate ACV bound at the active site, its cysteinyl β-carbon exposed to attack by a second molecule of substrate, adjacent. Formation of this product constitutes a new mode of reaction for IPNS and non-heme iron oxidases in general.

PubMed: 28703303
DOI: 10.1002/chem.201701592

Experimental method

X-RAY DIFFRACTION (1.3 Å)