Abstract
Brassinolide, the most biologically active brassinosteroid, is formed by a Baeyer-Villiger oxidation of castasterone. However, such enzyme has not been identified. The formation of castasterone from 6-deoxocastasterone via two-step oxidations is catalyzed by cytochrome P450 monooxygenases (CYP85As). A null mutation in the tomato CYP85A1 gene, extreme dwarf (dx), causes severe dwarfism due to brassinosteroid deficiency, but this mutant still produces normal fruits. GC-MS analysis showed that dx fruits, as well as the wild type, produce brassinolide at high levels, indicating that brassinolide is synthesized in the fruits independently of CYP85A1. We have identified a new CYP85A gene that is specifically expressed in tomato fruits. This newly identified CYP85A, transformed into yeast, catalyzed the Baeyer-Villiger oxidation of castasterone to brassinolide, as well as the conversion of 6-deoxocastasterone to castasterone. Thus, this enzyme has been identified with the brassinolide synthase of tomato which is a multifunctional P450 enzyme catalyzing three consecutive oxidations.
