Interactions of isopenicillin N synthase with cyclopropyl-containing substrate analogues reveal new mechanistic insight.
Isopenicillin N synthase (IPNS), a non-heme iron oxidase central to penicillin and cephalosporin biosynthesis, catalyzes an energetically demanding chemical transformation to produce isopenicillin N from the tripeptide delta-(l-alpha-aminoadipoyl)-l-cysteinyl-d-valine (ACV). We describe the synthesis of two cyclopropyl-containing tripeptide analogues, delta-(l-alpha-aminoadipoyl)-l-cysteinyl-beta-methyl-d-cyclopropylglycine and delta-(l-alpha-aminoadipoyl)-l-cysteinyl-d-cyclopropylglycine, designed as probes for the mechanism of IPNS. We have solved the X-ray crystal structures of these substrates in complex with IPNS and propose a revised mechanism for the IPNS-mediated turnover of these compounds. Relative to the previously determined IPNS-Fe(II)-ACV structure, key differences exist in substrate orientation and water occupancy, which allow for an explanation of the differences in reactivity of these substrates.
Howard-Jones
Annaleise R
AR
Chemistry Research Laboratory, University of Oxford, Mansfield Road, Oxford OX1 3TA, UK.
Elkins
Jonathan M
JM
Clifton
Ian J
IJ
Roach
Peter L
PL
Adlington
Robert M
RM
Baldwin
Jack E
JE
Rutledge
Peter J
PJ
D001233:Aspergillus nidulans / Q000201:enzymology; D003460:Crystallization; D018360:Crystallography, X-Ray; D008958:Models, Molecular; D009842:Oligopeptides / Q000138:chemical synthesis / Q000378:metabolism*; D010088:Oxidoreductases / Q000737:chemistry* / Q000378:metabolism
D009842:Oligopeptides; C520350:delta-(alpha-aminoadipoyl)-cysteinyl-beta-methylcyclopropylglycine; C520351:delta-(alpha-aminoadipoyl)-cysteinyl-cyclopropylglycine; D010088:Oxidoreductases; C038370:isopenicillin N synthetase