P-65: Effective Parameters on the Bovine Follicle Stimulating Hormone Expression in The Pichia Pastoris System

Background: Bovine follicle-stimulating hormone (bFSH) is a heterodimer hormone that consists of a common -subunit which noncovalently associated with the hormone-specific -subunit. During the past 15 years, the methylotrophic yeast Pichia pastoris has become an important host organism for recombinant protein production because it is able to use methanol as a sole carbon and energy source. The Pichia pastoris expression system is being used successfully for the production of various recombinant proteins such as FSH. Optimization of conditions during expression for improving the yield of protein production is a major goal for scientists. The aim of this project is to study the effect of temperature and methanol induction yield on the expression of bFSH in the methylotrophic yeast pichia pastoris. Materials and Methods: In this study, bovine FSH recombinant protein were produced in Pichia pastoris expression system using BMMY medium at pH=6, Temperatures of induction (15, 18, 21, 24, 27, 30, 32°C) and methanol concentration used for induction (0.5, 1, 2, 3%) in five day incubation time. The experiments were carried out in order to find the optimal temperature and methanol induction yield. The recombinant proteins were precipitate using TCA method. The protein evaluated by SDS-PAGE followed by Western blotting with specific antibody for βFSH and a ~16-kDa protein of interest was confirmed. Results: Results showed that temperature and methanol significantly affect on expression of bFSH. The best temperature for Expression of bovine FSH is 30°C and the optimal methanol induction yield for expression system is 1%. Conclusion: The findings demonstrate how temperature and methanol influences on production of recombinant protein bFSH in the methylotrophic yeast pichia pastoris.