Secreted expression of the VP2 protein of very virulent infectious bursal disease virus in the methylotrophic yeast Pichia pastoris

Abstract

The VP2-encoding gene of very virulent infectious bursa] disease virus (vvIBDV) was amplified using reverse transcription (RT)-polymerase chain reaction (PCR) and inserted into pPICZalphaA vector. Recombinant plasmid DNA was integrated into the chromosome of the transformed Pichia pastoris by electroporation and expressed protein identified by SDS-PAGE and Western blotting. High-level secreted expression was performed by determining the Mut(+) phenotype and secreting multi-copy integrants in the recombinant yeast. A recombinant protein of approximately 67 kDa was secreted into the supernatant from the yeast when induced with methanol. The expressed supernatant was bound with chicken anti-IBDV polyclonal antibodies. Western blotting with antibodies against vvIBDV indicated that the recombinant VP2 protein retained its antigenicity. High-level production (10mg/100m]) of the recombinant VP2 protein indicated that P pastoris was an efficent expression system for vvIBDV VP2 protein. (C) 2004 Elsevier B.V. All rights reserved.