Crystallographic studies on the binding of selectively deuterated LLD- and LLL-substrate epimers by isopenicillin N synthase |
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Authors: | Wei Ge Jeanette E Stok Robert M Adlington Peter J Rutledge |
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Institution: | a Chemistry Research Laboratory, University of Oxford, Mansfield Rd., Oxford OX1 3TA, UK b School of Chemistry F11, The University of Sydney, NSW 2006, Australia |
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Abstract: | Isopenicillin N synthase (IPNS) is a non-heme iron(II) oxidase which catalyses the biosynthesis of isopenicillin N (IPN) from the tripeptide δ-l-α-aminoadipoyl-l-cysteinyl-d-valine (lld-ACV). Herein we report crystallographic studies to investigate the binding of a truncated lll-substrate in the active site of IPNS. Two epimeric tripeptides have been prepared by solution phase peptide synthesis and crystallised with the enzyme. δ-l-α-Aminoadipoyl-l-cysteinyl-d-2-amino-3,3-dideuteriobutyrate (lld-ACd2Ab) has the same configuration as the natural substrate lld-ACV at each of its three stereocentres; its epimer δ-l-α-aminoadipoyl-l-cysteinyl-l-2-amino-3,3-dideuteriobutyrate (lll-ACd2Ab) has the opposite configuration at its third amino acid. lll-ACV has previously been shown to inhibit IPNS turnover of its substrate lld-ACV; the all-protiated tripeptide δ-l-α-aminoadipoyl-l-cysteinyl-d-2-aminobutyrate (lld-ACAb) is a substrate for IPNS, being turned over to a mixture of penam and cepham products. Comparisons between the crystal structures of the IPNS:Fe(II):lld-ACd2Ab and IPNS:Fe(II):lll-ACd2Ab complexes offer a possible rationale for the previously observed inhibitory effects of lll-ACV on IPNS activity. |
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Keywords: | AC6FV δ-α-aminoadipoyl-cysteinyl-3 3 3 3&prime 3&prime 3&prime -hexafluorovaline ACA δ-α-aminoadipoyl-cysteinyl-alanine ACd2Ab δ-α-aminoadipoyl-cysteinyl-2-amino-3 3-dideuteriobutyrate ACG δ-α-aminoadipoyl-cysteinyl-glycine ACOmC δ-α-aminoadipoyl-cysteine (1-carboxy-2-thiomethyl)ethyl ester ACV δ-α-aminoadipoyl-cysteinyl-valine IPN isopenicillin N IPNS isopenicillin N synthase NHIO non-heme iron(II) oxidase |
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