Expression of strictosidine P-D-glucosidase cDNA from Catharanthus roseus,involved in the monoterpene indole alkaloid pathway, in a transgenic suspension culture of Nicotiana tabacum

Different transgenic cell lines of Nicotiana tabacum expressing strictosidi ne beta -D-glucosidase cDNA from Catharanthus roseus were established follo wing Agrobacterium tumefaciens infection. The fresh weight and protein leve ls did not appear to be affected by the gene insertion performed, and in se veral instances these seemed to be boosted. Regarding strictosidine beta -D -glucosidase activity (SGD, EC 3.2.1.105) in some of the cell lines, this s eemed growth associated but in others it appeared steady during growth, sug gesting a continuous activation of the inserted gene cassette triggered by the CaM35S promoter, showing maximum values of ca. 170 pkat(.)mg(-1) protei n following incubation of crude protein extracts. Liquid nutrient medium fa iled to show any enzyme activity and lacked proteins, indicating that the g ene product was not released into the medium. Molecular data showed, as see n in C. roseus cells, that SGD activity was associated with a protein aggre gate of a size of 650 kDa, and this was absent in control and samples of th e transgenic lines which failed to show SGD activity. (C) 2001 Editions sci entifiques et medicales Elsevier SAS.