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.

译文

:我们研究了洗涤剂对拟南芥(CYP74A1)的氧化烯合酶(AOS)动力学和低聚状态的影响。我们显示不含洗涤剂的CYP74A1是单体的,具有双特异性的高度水溶性,但活性相对较低。洗涤剂微团促进k(cat)/ K(m)增加48倍(至5.9 x 10(7)M(-1)s(-1)),同时血红素铁的自旋态平衡发生变化由于单个去污剂胶束与蛋白质单体结合,这是非典型的P450酶。已显示该机制是CYP74A1底物特异性的重要决定因素。 CYP74A1可能适合第一种植物细胞色素P450的结构解析,其9-AOS活性和体外行为对其在植物中的作用具有影响。

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