Expression, purification and characterization of WSSV-VP37 in Pichia pastoris

Abstract

White spot syndrome virus (WSSV) is one of the major pathogens in cultured shrimp. However, up to now, the mechanisms of virus attachment and entry into shrimp cells are poorly understood. Recent research has indicated that WSSV-VP37 is one of the virus attachment proteins of WSSV. Therefore it is important to express the WSSV-VP37 gene in order to determine how it binds to the shrimp cells and interacts in the shrimp. In this study, Pichia pastoris was used as a host to express the WSSV-VP37 gene. The VP37 gene was amplified from the genomic DNA of a Chinese isolate of WSSV and cloned in P. pastoris expression vector pGAPZa-A. SDS-PAGE and Western blot analyses showed that VP37 migrated as a single band with a molecular weight of 37 kDa, and that mouse anti-(His)×6-antibodies bound specifically to VP37. The culture supernatant of the VP37 expression strain, SMD1168/pGAPZaA-VP37, was purified to greater than 95% homogeneity using ammonium sulfate precipitation and an Ni–NTA column. Binding assay by ELISA and fluorescence microscopy in vitro indicated that VP37 is capable of binding to shrimp cells. These results indicate that the P. pastoris expression system is useful for production of bioactive WSSV-VP37 and that WSSV-VP37 is an attachment protein in the infection of shrimp.