Monooxygenases involved in GA(12) and GA(14) synthesis in Gibberella fujikuroi

A microsomal preparation from mycelia of the gibberellin (GA)-producing fun gus Gibberella fujikuroi catalyzed the first two steps in the conversion of the biosynthetic intermediate GA(12)-aldehyde to gibberellic acid (GA(3)). [C-14]GA(12)-Aldehyde was converted to radiolabelled GA(14), the major pro duct, together with smaller amounts of non-hydroxylated GA(12). The microso mal activities required reduced pyridine nucleotides and molecular oxygen. However, GA(12) and GA(14) synthesis differed markedly in the preferred ele ctron source. Formation of GA(12) required NADH or NADPH, while GA(14) synt hesis from GA(12)-aldehyde occurred only with NADPH. Marked differences wer e also found in the activating effect of FAD. When NADPH was the reductant, the rate of GA(14) synthesis was enhanced 3.5 times by 5 muM FAD while thi s flavin nucleotide did not alter the synthesis of GA(12). In contrast, GA( 12) synthesis was activated 3.5 times by 50 muM FAD in the presence of NADH . Both activities were inhibited by carbon monoxide and cytochrome c. These properties suggest that the 3 beta -hydroxylation of GA(12)-aldehyde and f urther oxidation of carbon 7 are catalyzed by cytochrome P-450 monooxygenas es in Gibberella fujkuroi . (C) 2001 Elsevier Science Ltd. All rights reser ved.