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Expression and purification of recombinant human serum albumin fusion protein with VEGF165b in Pichia pastoris.


Protein Expr Purif. 2012 Jun 27;


Authors: Zhu RY, Xin X, Dai HY, Li Q, Lei JY, Chen Y, Jin J


Abstract

VEGF165b is an endogenous transcriptional splice variant of VEGF and has been shown to have a therapeutic potency as an anti-cancer agent. In this report, a fusion gene consisting of a human VEGF165b and a human albumin (HSA) gene was constructed and then inserted into a pPIC9k vector. The recombinant fusion protein, rhHSA-VEGF165b, was over expressed in the methylotrophic yeast Pichia pastoris under the control of AOX1 promoter. After induction with methanol, the expression level of rhHSA-VEGF165b was 275mg/L in broth. The fusion protein rhHSA-VEGF165b was purified to more than 95% purity by using Blue Sepharose Fast Flow and SP Sepharose Fast Flow. Biological activity of the prepared rhHSA-VEGF165b was characterized by transwell migration assay, retaining about 9% of that of unmodified rhVEGF165b on a molar basis. Data from mice show that the serum half-life time of rhHSA-VEGF165b was nearly 20 times longer than that of rhVEGF165b.

PMID: 22750397 [PubMed - as supplied by publisher]