Expression Of HBmp4 In Pichia Pastoris And The Effect Of Codon Optimization Of HBmp4 On The Expression

Bone morphogenetic proteins ( BMPs ) are dimeric secreted glycoproteins, each members shares sequence similarity in their C'terminal regions. BMPs have diverse biological activities and play critical roles in the migration, proliferation, and differentian of mesenchymal cells during embryogenesis and in the repair and regeneration of tissues during postfetal life. In our research, the recombinant human bone morphogenetic proteins 4 ( rhBMP4 ) was sussessfully expressed in the Pichia pastoris. Furthermore, investigations for rhBMP4 expression, small scale fermentations were carried out.The cDNA mature domain encoding hBMP4 was cloned into the expression vector, pPIC9K. And the reconstructed Pichia pastoris expression vector was introduced into Pichia pastoris strain for expression. For higher expression level, codons in the cDNA mature domain encoding hBMP4 were optimized according to the favorite for Pichia pastoris. Analysis of culture medium revealed that the molecular weight of rhBMP4 is about 26KD by SDS-PAGE and WESTERN-BLOT; the yield is about 17.731mg/L and accounts for 22.115% of the total proteins in the supernatants. Expression with favorite codons for Pichia pastoris increased about 3-fold in proportion to that with the original codons. The optimal time to harvest is 96h post induction with 0.5% of methanol, with the culture medium buffered at pH6.0 and the optimal OD₆00 at about 10 before induction.