Ethylene and Methyl Jasmonate Interaction and Binding Models For Elicited Biosynthetic Steps of Paclitaxel in Suspension Cultures of Taxus Canadensis

Abstract

Ethylene and methyl jasmonate (MJ) act as co-mediators of defined cellular responses in many plant systems. We postulate that specific biosynthetic steps for taxane production are regulated by allosteric regulation of ethylene binding, based on our absorption and induction modeling of paclitaxel formation in elicited suspension cell cultures of Taxus canadensis. Binding sites for ethylene on cellular membranes from many plant genera have been established for years, and recently the transmembrane ethylene receptor protein, ETR1, has been characterized as a two-component regulator. In attempting to model this system, we view a two-step process: MJ absorption in the membrane is directly related to MJ concentration, but its interaction with ETR1 is effective only at higher concentrations. Hence, at low MJ concentrations (0-20 μM), the unmodulated ethylene binding blocks induction of enzymes that either synthesize 3-phenylisoserine and/or add the sidechain to the baccatin III ring. Sigmoidal plots of paclitaxel and 7-xylosyl-10-deacetyltaxol productivity versus MJ concentration indicates allosteric modulation of ethylene binding by MJ. At medium to high MJ concentrations (200-400 μM), the modulation site is saturated, and no greater productivities are seen. These data logically fit an induction model described by Mirjalili and Linden [22]. Dependence on ethylene concentration is observed (27>10>7>3 μL/L) in baccatin III accumulation, but 7-xylosyl-10-deacetyltaxol and taxol in the culture medium are maximized using continuous 7 μL/L headspace (estimated 70 nM dissolved) ethylene, that is supplied independently to replicated shake flasks using flow along with uniform 10 (v/v) % oxygen and 0.5 (v/v) % carbon dioxide (balance nitrogen) from a gas mixing station.