Tesis doctoral de David Resina Rodriguez
During the course of this work, the expression of a rhizopus oryze lipase (rol) under the control of the fld1 promoter in the methylotrophic yeast pichia pastoris, as well as the development of methanol-free cultivation processes and metabolic engineering strategies for process improvement has been carried out. first, an expression vector containing the rol gene and the fld1 promoter from p. Pastoris was constructed and subsequently transformed in p. Pastoris. Subsequent studies in shake flask and bioreactor batch cultivations revealed the synergetic effect of methanol and methylamine on the fld1 promoter induction. Moreover, the use of sorbitol as carbon source, combined with methylamine as nitrogen source, was found to be an attractive alternative to the use of methanol for high cell density cultivations. second, a new fed-batch fermentation strategy was developed using sorbitol and methylamine as carbon and nitrogen source, respectively. Three fed-batches were performed at different specific growth rates, 0.005 h-1, 0.01 h-1, and 0.02 h-1. The specific growth rate was controlled during induction phase in the 0.005 h-1, 0.01 h-1 fermentations by implementing an exponential feeding rate at limiting substrate concentration. In the near-to-maximum specific growth rate fermentation performed at about 0.02 h-1, sorbitol concentration was keep into the medium at about 8 g·l-1 throughout the induction phase. The highest lipase productivity was obtained in the fed-batch performed at the 0.02 h-1 specific growth rate. However, product production rate of all three fermentations dropped soon after reaching a maximum at about 30 hours of culture. third, in order to analyze potential bottlenecks in the protein folding or secretion pathway and, in particular, the potential activation of the unfolded protein response (upr) during rol overexpression and intracellular accumulation, the levels of a reticle endoplasmatic-resident chaperone (bip), its induction has been relate
Datos académicos de la tesis doctoral «Expression of the rhizopus oryzae lipase in pichia pastoris under the control of the fld1 promoter«
- Título de la tesis: Expression of the rhizopus oryzae lipase in pichia pastoris under the control of the fld1 promoter
- Autor: David Resina Rodriguez
- Universidad: Autónoma de barcelona
- Fecha de lectura de la tesis: 30/09/2006
Dirección y tribunal
- Director de la tesis
- Francisco Valero Barranco
- Tribunal
- Presidente del tribunal: josep Lopez santin
- daniel Ramón vidal (vocal)
- joaquin Ariño (vocal)
- xavier Berthet (vocal)