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Human beta-nerve growth factor expressed in yeast by fusion to ubiquitin Garcia, Julia N.; Ubieta, Raimundo; Seralena, Alina; Santos, Alicia; Dominguez, Maria del C.; Jimenez, Victor; de las M. Vazquez, Maria; Garcia, Jose & Menendez, Alfredo Abstract An expression system has been established in Pichia pastoris to yield recombinant human-Nerve Growth Factor where the C-terminus of ubiquitin protein was fused to the N-terminus of beta-NGF. The human Nerve Growth Factor gene was integrated into the chromosome of the P. pastoris under the control of the methanol-regulated alcohol oxidase promoter and fusioned to the ubiquitin gene. The fusion protein was expressed and processed in vivo. An ubiquitin C-terminal hydrolase cleaves the junction peptide bond between the ubiquitin and beta-NGF. The identity of NGF was demonstrated by Western blot. Keywords Recombinant nerve growth factor, ubiquitin, P. pastoris

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Garcia, Julia N.; Ubieta, Raimundo; Seralena, Alina; Santos, Alicia; Dominguez, Maria del C.; Jimenez, Victor; de las M. Vazquez, Maria; Garcia, Jose & Menendez, Alfredo Resumen Un sistema de expresion ha sido establecido en Pichia pastoris para producir NGF humano recombinante, donde el extremo C-terminal de la ubiquitina se fusiono al extremo N-terminal del beta-NGF. El gen de NGF se integro en el cromosoma de Pichia pastoris bajo el control del promotor de la enzima alcohol oxidasa, el cual es fuertemente regulado por metanol. Esta proteina de fusion se expreso y proceso in vivo. Una enzima hidrolasa del sistema ubiquitina corta la fusion. Se demostro por Western blot la identidad del NGF

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