Structure and peroxidase activity of ferric Streptomyces clavuligerus orf10-encoded protein P450CLA: UV-visible, CD, MCD and EPR spectroscopic characterization

Journal of the Brazilian Chemical Society

Leandro S. Goto²  Carlos O. Hokka²  José F. Lima¹  Tatiana Prieto¹  Ana Paula U. Araújo¹  Iseli L. Nantes¹  Otaciro R. Nascimento¹ 

[1] ,Universidade Federal de São Carlos Departamento de Engenharia Química Grupo de Engenharia BioquímicaSão Carlos SP ,Brazil

关键词: cytochrome P450;    orf10;    peroxidase activity;    magnetic circular dichroism;    electron paramagnetic resonance;    spin trapping;   

DOI  :  10.1590/S0103-50532012000500017

来源: SciELO

【 摘 要 】

The present study reports the spectroscopic characterization by UV-visible absorption spectroscopy, magnetic circular dichroism (MCD) and electron paramagnetic resonance (EPR) of the recombinant orf10-encoded P450-camphor like protein (P450CLA)of Streptomyces clavuligerus expressed in Escherichia coli Rosetta in the native form and associated to external ligands containing the β-lactam, oxazole and alkylamine-derived (alcohol) moieties of the clavulamic acid. Considering the diversity of potential applications for the enzyme, the reactivity with tert-butylhydroperoxide (tert-BuOOH) was also characterized. P450CLA presents a covalently bound heme group and exhibited the UV-visible, CD and MCD spectral features of P450CAM including the fingerprint Soret band at 450 nm generated by the ferrous CO-complex. P450CLA was converted to high valence species by tert-BuOOH and promoted homolytic scission of the O-O bond. The radical profile of the reaction was tert-butyloxyl as primary and methyl and butylperoxyl as secondary radicals. The secondary methyl and butylperoxyl radicals resulted respectively from the β-scission of the alkoxyl radical and from the reaction of methyl radical with molecular oxygen.

【 授权许可】

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