Amorpha-4,11-diene synthase of Artemisia annua: cDNA isolation and bacterial expression of a terpene synthase involved in artemisinin biosynthesis

作者：Chang YJ; Song SH; Park SH; Kim SU^*

来源：Archives of Biochemistry and Biophysics, 2000, 383(2): 178-184.

DOI：10.1006/abbi.2000.2061

• amorpha-4,11-diene synthase
• sesquiterpene synthase
• rapid amplification of cDNA ends
• artemisinin biosynthesis
• Artemisia annua

Artemisia annua, an indigenous plant to Korea, contains an antimalarial sesquiterpene, artemisinin. The first committed step of artemisinin biosynthesis is the cyclization of farnesyl diphosphate by a sesquiterpene synthase to produce an amorphane-type ring system. The aims of this research were to molecularly clone and express amorpha-4,11-diene synthase for metabolic engineering. PCR amplification of genomic DNA with a pair of primers, designed from the conserved regions of sesquiterpene synthases of several plants, produced a 184-bp DNA fragment. This fragment was used in Northern blot analysis as a probe, showing approximately 2.2 kb of a single band. Its sequence information was used to produce 2106 bp of a full-length cDNA sequence including 1641 bp of open reading frame for 546 amino acids (kcs12) through a rapid amplification of cDNA ends (RACE). The deduced amino acid sequence displayed 36% identity with 5-epi-aristolochene synthase of Nicotiana tabacum. A soluble fraction of Escherichia coli harboring kcs12 catalyzed the cyclization of farnesyl diphosphate to produce a sesquiterpene, which was identified through GC-MS analysis as amorpha-4,11-diene.

• 出版日期2000-11-15