The P450–1 gene of Gibberella fujikuroi encodes a multifunctional enzyme in gibberellin biosynthesis

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Abstract
Recent studies have shown that the genes of the gibberellin (GA) biosynthesis pathway in the fungus Gibberella fujikuroi are organized in a cluster of at least seven genes. P450–1 is one of four cytochrome P450 monooxygenase genes in this cluster. Disruption of the P450–1 gene in the GA-producing wild-type strain IMI 58289 led to total loss of GA production. Analysis of the P450–1 -disrupted mutants indicated that GA biosynthesis was blocked immediately after ent -kaurenoic acid. The function of the P450–1 gene product was investigated further by inserting the gene into mutants of G. fujikuroi that lack the entire GA gene cluster; the gene was highly expressed under GA production conditions in the absence of the other GA-biosynthesis genes. Cultures of transformants containing P450–1 converted ent -[ 14 C]kaurenoic acid efficiently into [ 14 C]GA 14 , indicating that P450–1 catalyzes four sequential steps in the GA-biosynthetic pathway: 7β-hydroxylation, contraction of ring B by oxidation at C-6, 3β-hydroxylation, and oxidation at C-7. The GA precursors ent- 7α-hydroxy[ 14 C]kaurenoic acid, [ 14 C]GA 12 -aldehyde, and [ 14 C]GA 12 were also converted to [ 14 C]GA 14 . In addition, there is an indication that P450–1 may also be involved in the formation of the kaurenolides and fujenoic acids, which are by-products of GA biosynthesis in G. fujikuroi . Thus, P450–1 displays remarkable multifunctionality and may be responsible for the formation of 12 products.

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