Enzymes in ginkgolide biosynthetic pathway : cloning and genomic structure

Abstract

Part 1 The Ginkgo biloba is a perennial woody plant growing throughout the whole region of Korea. It accumulates various ginkgolides, whose oxidative modification step of terpenoid intermediates in biosynthesis is monooxygenation by cytochrome P450 monooxygenases. The partial cDNAs from G. biloba root using homology-based PCR yielded five distinct cytochrome P450 families (GbCYP∙A, GbCYP∙B, GbCYP∙C, GbCYP∙D, GbCYP∙E). The GbCYP∙E sequence information derived from rapid amplification of cDNA ends (RACE) was used to produce 1655 bp of a full-length cDNA sequence including 1455 bp of an open reading frame for 485 amino acids with a deduced size of 55.172 kDa. The deduced amino acid sequence displayed 29% identity with taxane 14β-hydroxylase of Taxus cuspidata. Also, a cytochrome P450 cDNA which was 2038 bplong and partially deficient of the 5 -UTR (untranslated region) has been isolated from the cDNA library of G. biloba root, naming it as GbCYP∙F. This showed to have an open reading frame 1761 bp-long and a 586 amino acid sequence with its size 66.196 kDa. The deduced amino acid sequences of GbCYP∙F displayed 75% identity with CYP97B2 monooxygenase of Glycine max. This is the first report on cDNA isolation of the cytochrome P450 genes from G. biloba.