Expression of a Rice Thaumatin-Like Protein (PR-5) Gene in a Baculovirus Expression System

We previously isolated a full-length thaumatin-like protein (TLP) cDNA clone (D34) from a rice cDNA library prepared from RNA isolated from R. solani-infected rice plants. Furthermore, it has been demonstrated that transgenic rice plants that are constitutively expressing the TLP (D34) show enhanced resistance to R. solani. In order to purify and characterize this TLP. we expressed it (D34) in Spodoptera frugiperda (sf21) insect cells using a baculovirus expression system. Western blot analysis indicated that a 23-kDa protein cross-reacting with a bean TLP antibody was produced abundantly in the insect cells. However, this protein was not secreted into the culture medium. The N-terminal sequence of the protein (19 residues) showed high sequence similarity to thaumalin and TLPs of oat, barley, maize, grapes and tomato. The cleavage site for the TLP was determined as immediately upstream of the sequence ATFAITN.