title: Allene oxide synthase from Arabidopsis thaliana (CYP74A1) exhibits dual specificity that is regulated by monomer-micelle association.
abstract: We investigate the effects of detergent on the kinetics and oligomeric state of allene oxide synthase (AOS) from Arabidopsis thaliana (CYP74A1). We show that detergent-free CYP74A1 is monomeric and highly water soluble with dual specificity, but has relatively low activity. Detergent micelles promote a 48-fold increase in k(cat)/K(m) (to 5.9 x 10(7)M(-1)s(-1)) with concomitant changes in the spin state equilibrium of the haem-iron due to the binding of a single detergent micelle to the protein monomer, which is atypical of P450 enzymes. This mechanism is shown to be an important determinant of the substrate specificity of CYP74A1. CYP74A1 may be suited for structural resolution of the first plant cytochrome P450 and its 9-AOS activity and behaviour in vitro has implications for its role in planta.
authors.affiliation: John Innes Centre, Norwich Research Park, Norwich NR4 7UH, United Kingdom. Richard.Hughes@bbsrc.ac.uk
authors.forename: Richard K
authors.initials: RK
authors.lastname: Hughes
authors.forename: Eric J
authors.initials: EJ
authors.lastname: Belfield
authors.forename: Ruth
authors.initials: R
authors.lastname: Ashton
authors.forename: Shirley A
authors.initials: SA
authors.lastname: Fairhurst
authors.forename: Cornelia
authors.initials: C
authors.lastname: Göbel
authors.forename: Michael
authors.initials: M
authors.lastname: Stumpe
authors.forename: Ivo
authors.initials: I
authors.lastname: Feussner
authors.forename: Rod
authors.initials: R
authors.lastname: Casey
mesh_terms: D017360:Arabidopsis / Q000201:enzymology*; D029681:Arabidopsis Proteins / Q000737:chemistry*; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry*; D003902:Detergents / Q000737:chemistry; D019746:Intramolecular Oxidoreductases; D007700:Kinetics; D008823:Micelles; D013379:Substrate Specificity
chemical_list: D029681:Arabidopsis Proteins; D003902:Detergents; D008823:Micelles; D003577:Cytochrome P-450 Enzyme System; D019746:Intramolecular Oxidoreductases; C021377:hydroperoxide isomerase