The crystal structure of an LLL-configured depsipeptide substrate analogue bound to isopenicillin N synthase.
Isopenicillin N synthase (IPNS) is a non-heme iron(ii) oxidase, which catalyses the biosynthesis of isopenicillin N (IPN) from the tripeptide delta-l-alpha-aminoadipoyl-l-cysteinyl-d-valine (lld-ACV) in a remarkable oxidative bicyclisation reaction. The natural substrate for IPNS is the lld-configured tripeptide. lll-ACV is not turned over by the enzyme, but inhibits turnover of the lld-tripeptide. The mechanism by which this inhibition takes place is not fully understood. Recent studies have employed a range of lld-configured depsipeptide substrate analogues in crystallographic studies to probe events preceding beta-lactam closure in the IPNS reaction cycle. Herein, we report the first crystal structure of IPNS in complex with an lll-configured depsipeptide analogue, delta-l-alpha-aminoadipoyl-l-cysteine (1-(R)-carboxy-2-thiomethyl)ethyl ester (lll-ACOmC). This report describes the crystal structure of the IPNS:Fe(ii):lll-ACOmC complex to 2.0 A resolution, and discusses attempts to oxygenate this complex at high pressure in order to probe the mechanism by which lll-configured substrates inhibit IPNS catalysis.
Ge
Wei
W
Chemistry Research Laboratory, University of Oxford, UK.
Clifton
Ian J
IJ
Stok
Jeanette E
JE
Adlington
Robert M
RM
Baldwin
Jack E
JE
Rutledge
Peter J
PJ
D001233:Aspergillus nidulans / Q000737:chemistry* / Q000378:metabolism; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D047630:Depsipeptides / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D015394:Molecular Structure; D010088:Oxidoreductases / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D013379:Substrate Specificity
D047630:Depsipeptides; D010088:Oxidoreductases; C038370:isopenicillin N synthetase