Improved expression of fusion protein using a cysteine- protease- and chitinase-deficient Bombyx mori (silkworm) multiple nucleopolyhedrovirus bacmid in silkworm larvae.

A bacmid system of BmMNPV [Bombyx mori (silkworm) multiple nucleopolyhedrovirus] with both the cysteine-protease and chitinase genes deleted (BmMNPV-CP(-)-Chi(-)) was constructed. The viral protease and chitinase activities in the haemolymph of Bombyx mori larvae infected with this BmMNPV-CP(-)-Chi(-) bacmid were reduced by 95 and 50% respectively. By using this system, a GFP(uv)-beta3GnT2 (green fluorescent protein excited by UV light-beta1,3-N-acetylglucosaminyltransferase 2) fusion protein was successfully expressed in silkworm larvae with less protein degradation and without larvae liquefaction; beta3GnT2 activity improved 2.8-fold over that of unmodified bacmid. This BmMNPV-CP(-)-Chi(-) bacmid system provides rapid protein production in silkworms and can be used for the production of recombinant eukaryotic proteins without proteolytic degradation.