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PMID: 20797613 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S.

Structure of cytochrome P450 PimD suggests epoxidation of the polyene macrolide pimaricin occurs via a hydroperoxoferric intermediate.

Chemistry & biology ·Vol. 17 ·No. 8 ·2010-08-27 ·Pages 841-51

Kells PM, Ouellet H, Santos-Aberturas J, Aparicio JF, Podust LM

Abstract

We present the X-ray structure of PimD, both substrate-free and in complex with 4,5-desepoxypimaricin. PimD is a cytochrome P450 monooxygenase with native epoxidase activity that is critical in the biosynthesis of the polyene macrolide antibiotic pimaricin. Intervention in this secondary metabolic pathway could advance the development of drugs with improved pharmacologic properties. Epoxidation by P450 typically includes formation of a charge-transfer complex between an oxoferryl pi-cation radical species (Compound I) and the olefin pi-bond as the initial intermediate. Catalytic and structural evidence presented here suggest that epoxidation of 4,5-desepoxypimaricin proceeds via a hydroperoxoferric intermediate (Compound 0). The oxygen atom of Compound 0 distal to the heme iron may insert into the double bond of the substrate to make an epoxide ring. Stereoelectronic features of the putative transition state suggest substrate-assisted proton delivery.

MeSH Terms
Amino Acid Sequence Anti-Bacterial Agents/chemistry,metabolism Benzene/chemistry,metabolism Biocatalysis Crystallography, X-Ray Cytochrome P-450 Enzyme System/chemistry,metabolism Epoxy Compounds/metabolism Models, Molecular Molecular Sequence Data Natamycin/chemistry,metabolism Peroxynitrous Acid/metabolism Protein Conformation Protons Stereoisomerism Substrate Specificity
Chemicals
Anti-Bacterial Agents Epoxy Compounds Protons Peroxynitrous Acid Natamycin Cytochrome P-450 Enzyme System Benzene
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Kells Petrea M
Department of Pharmaceutical Chemistry, University of California, San Francisco, CA 94158, USA.
Ouellet Hugues
Santos-Aberturas Javier
Aparicio Jesus F
Podust Larissa M
References (49)
49 references, click to expand
  1. Heme-Containing Oxygenases.
    Chem Rev. 1996 Nov 7;96(7):2841-2888 PMID: 11848843
  2. Polyene macrolide antibiotic biosynthesis.
    Curr Med Chem. 2004 Jun;11(12):1645-56 PMID: 15180569
  3. ESPript: analysis of multiple sequence alignments in PostScript.
    Bioinformatics. 1999 Apr;15(4):305-8 PMID: 10320398
  4. Engineered biosynthesis of novel polyenes: a pimaricin derivative produced by targeted gene disruption in Streptomyces natalensis.
    Chem Biol. 2001 Jul;8(7):635-44 PMID: 11451665
  5. Investigation of the proton-assisted pathway to formation of the catalytically active, ferryl species of P450s by molecular dynamics studies of P450eryF.
    J Am Chem Soc. 1996 Jul 10;118(27):6377-87 PMID: 11540056
  6. Characterization of the polyene macrolide P450 epoxidase from Streptomyces natalensis that converts de-epoxypimaricin into pimaricin.
    Biochem J. 2005 Feb 15;386(Pt 1):57-62 PMID: 15228385
  7. Rapid kinetics investigations of peracid oxidation of ferric cytochrome P450cam: nature and possible function of compound ES.
    J Inorg Biochem. 2006 Dec;100(12):2034-44 PMID: 17095096
  8. Quantitative structure-activity relationships in amphotericin B derivatives.
    Biochem Pharmacol. 1988 Mar 1;37(5):827-36 PMID: 3278711
  9. On the identity and reactivity patterns of the "second oxidant" of the T252A mutant of cytochrome P450cam in the oxidation of 5-methylenenylcamphor.
    J Inorg Biochem. 2006 Dec;100(12):2054-68 PMID: 17084458
  10. Kinetic characterization of compound I formation in the thermostable cytochrome P450 CYP119.
    J Biol Chem. 2002 Mar 22;277(12):9641-4 PMID: 11799104
  11. The CCP4 suite: programs for protein crystallography.
    Acta Crystallogr D Biol Crystallogr. 1994 Sep 1;50(Pt 5):760-3 PMID: 15299374
  12. Multiple mechanisms and multiple oxidants in P450-catalyzed hydroxylations.
    Arch Biochem Biophys. 2003 Jan 1;409(1):72-9 PMID: 12464246
  13. Uncoupling of the cytochrome P-450cam monooxygenase reaction by a single mutation, threonine-252 to alanine or valine: possible role of the hydroxy amino acid in oxygen activation.
    Proc Natl Acad Sci U S A. 1989 Oct;86(20):7823-7 PMID: 2510153
  14. Polyene macrolide antibiotics and their applications in human therapy.
    Curr Med Chem. 2003 Feb;10(3):211-23 PMID: 12570708
  15. Biosynthetic engineering of polyene macrolides towards generation of improved antifungal and antiparasitic agents.
    Curr Top Med Chem. 2008;8(8):639-53 PMID: 18473889
  16. Hydroxylation by the hydroperoxy-iron species in cytochrome P450 enzymes.
    J Am Chem Soc. 2004 Jan 14;126(1):115-26 PMID: 14709076
  17. Crystal structures of the ferrous dioxygen complex of wild-type cytochrome P450eryF and its mutants, A245S and A245T: investigation of the proton transfer system in P450eryF.
    J Biol Chem. 2005 Jun 10;280(23):22102-7 PMID: 15824115
  18. Epoxidation of olefins by hydroperoxo-ferric cytochrome P450.
    J Am Chem Soc. 2003 Mar 26;125(12):3406-7 PMID: 12643683
  19. Refinement of macromolecular structures by the maximum-likelihood method.
    Acta Crystallogr D Biol Crystallogr. 1997 May 1;53(Pt 3):240-55 PMID: 15299926
  20. Improved antifungal polyene macrolides via engineering of the nystatin biosynthetic genes in Streptomyces noursei.
    Chem Biol. 2008 Nov 24;15(11):1198-206 PMID: 19022180
  21. Spectroscopic investigations of intermediates in the reaction of cytochrome P450(BM3)-F87G with surrogate oxygen atom donors.
    J Inorg Biochem. 2006 Dec;100(12):2045-53 PMID: 17083977
  22. Cytochrome P450 oxidations in the generation of reactive electrophiles: epoxidation and related reactions.
    Arch Biochem Biophys. 2003 Jan 1;409(1):59-71 PMID: 12464245
  23. Biosynthesis of deoxyamphotericins and deoxyamphoteronolides by engineered strains of Streptomyces nodosus.
    Chem Biol. 2003 Dec;10(12):1215-24 PMID: 14700629
  24. Automated protein crystal structure determination using ELVES.
    Proc Natl Acad Sci U S A. 2004 Feb 10;101(6):1537-42 PMID: 14752198
  25. Strain energy of small ring hydrocarbons. Influence of C-h bond dissociation energies.
    J Am Chem Soc. 2004 Apr 7;126(13):4444-52 PMID: 15053635
  26. Hydroperoxoferric heme intermediate as a second electrophilic oxidant in cytochrome P450-catalyzed reactions.
    J Biol Inorg Chem. 2004 Sep;9(6):644-53 PMID: 15365901
  27. Polyene antibiotic biosynthesis gene clusters.
    Appl Microbiol Biotechnol. 2003 May;61(3):179-88 PMID: 12698274
  28. Crystal structure of CYP105A1 (P450SU-1) in complex with 1alpha,25-dihydroxyvitamin D3.
    Biochemistry. 2008 Apr 1;47(13):4017-27 PMID: 18314962
  29. Epoxidation of olefins by cytochrome P450: evidence from site-specific mutagenesis for hydroperoxo-iron as an electrophilic oxidant.
    Proc Natl Acad Sci U S A. 1998 Mar 31;95(7):3555-60 PMID: 9520404
  30. Molecular modelling of membrane activity of amphotericin B, a polyene macrolide antifungal antibiotic.
    Acta Biochim Pol. 2005;52(3):655-8 PMID: 16086075
  31. The Buccaneer software for automated model building. 1. Tracing protein chains.
    Acta Crystallogr D Biol Crystallogr. 2006 Sep;62(Pt 9):1002-11 PMID: 16929101
  32. Cytochrome p450 compound I.
    J Am Chem Soc. 2006 Apr 12;128(14):4580-1 PMID: 16594688
  33. Reactivity patterns of cytochrome P450 enzymes: multifunctionality of the active species, and the two states-two oxidants conundrum.
    Nat Prod Rep. 2007 Jun;24(3):533-52 PMID: 17534529
  34. Polyene macrolide antibiotic amphotericin B. Crystal structure of the N-iodoacetyl derivative.
    J Am Chem Soc. 1971 Sep 8;93(18):4560-4 PMID: 5131155
  35. Coot: model-building tools for molecular graphics.
    Acta Crystallogr D Biol Crystallogr. 2004 Dec;60(Pt 12 Pt 1):2126-32 PMID: 15572765
  36. Searching for the second oxidant in the catalytic cycle of cytochrome P450: a theoretical investigation of the iron(III)-hydroperoxo species and its epoxidation pathways.
    J Am Chem Soc. 2002 Mar 20;124(11):2806-17 PMID: 11890833
  37. Amphotericin B: current understanding of mechanisms of action.
    Antimicrob Agents Chemother. 1990 Feb;34(2):183-8 PMID: 2183713
  38. Replacement of tyrosine residues by phenylalanine in cytochrome P450cam alters the formation of Cpd II-like species in reactions with artificial oxidants.
    J Biol Inorg Chem. 2008 May;13(4):599-611 PMID: 18273651
  39. X-ray absorption spectroscopic characterization of a cytochrome P450 compound II derivative.
    Proc Natl Acad Sci U S A. 2008 Jun 17;105(24):8179-84 PMID: 18174331
  40. Reaction of ferric cytochrome P450cam with peracids: kinetic characterization of intermediates on the reaction pathway.
    J Biol Chem. 2005 May 27;280(21):20300-9 PMID: 15781454
  41. Crystal structures of epothilone D-bound, epothilone B-bound, and substrate-free forms of cytochrome P450epoK.
    J Biol Chem. 2003 Nov 7;278(45):44886-93 PMID: 12933799
  42. Spectra and kinetic studies of the compound I derivative of cytochrome P450 119.
    J Am Chem Soc. 2008 Oct 8;130(40):13310-20 PMID: 18788736
  43. Solution NMR structure of five representative glycosylated polyene macrolide antibiotics with a sterol-dependent antifungal activity.
    Eur J Biochem. 2002 Sep;269(18):4533-41 PMID: 12230565
  44. Substrate-assisted catalysis in cytochrome P450eryF.
    Nat Struct Biol. 1996 Jul;3(7):632-7 PMID: 8673608
  45. CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice.
    Nucleic Acids Res. 1994 Nov 11;22(22):4673-80 PMID: 7984417
  46. Oxidizing species in the mechanism of cytochrome P450.
    Nat Prod Rep. 2002 Aug;19(4):477-93 PMID: 12195813
  47. Interaction of amphotericin B and its selected derivatives with membranes: molecular modeling studies.
    Chem Rec. 2006;6(6):320-32 PMID: 17304519
  48. Characterization of the P450 monooxygenase NysL, responsible for C-10 hydroxylation during biosynthesis of the polyene macrolide antibiotic nystatin in Streptomyces noursei.
    Appl Environ Microbiol. 2006 Apr;72(4):2514-9 PMID: 16597951
  49. Mechanism of oxidation reactions catalyzed by cytochrome p450 enzymes.
    Chem Rev. 2004 Sep;104(9):3947-80 PMID: 15352783
Article Info
Journal
Chemistry & biology
Abbr.
Chem Biol
ISSN
1879-1301
DOI
10.1016/j.chembiol.2010.05.026
Published
2010-08-27
Pages
841-51
Language
English
Region
United States
NLM ID
9500160
PMCID
PMC2932657
Subset
IM
Grants
NIGMS NIH HHS · R01 GM078553-04 · United States
NIGMS NIH HHS · R01GM078553 · United States
NIGMS NIH HHS · R01 GM025515 · United States
NIGMS NIH HHS · R01 GM25515 · United States
NIGMS NIH HHS · R01 GM025515-33 · United States
NIGMS NIH HHS · R01 GM078553 · United States
NIGMS NIH HHS · R37 GM025515 · United States
Databases
PDB
2X9P, 2XBK
External Links
PubMed source DOI: 10.1016/j.chembiol.2010.05.026 PMC Full Text
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