Chitinase is responsible for insect chitin hydrolyzation, which is a key process in insect molting and pupation. However, little is known about the chitinase of Spodoptera exigua (SeChi). In this study, based on the SeChi gene (ADI24346) identified in our laboratory, we constructed the recombinant baculovirus P-Chi for the expression of recombinant SeChi (rSeChi) in Hi5 cells. The rSeChi was purified by chelate affinity chromatography, and the purified protein showed activity comparable with that of a commercial SgChi, suggesting that we harvested active SeChi for the first time. The purified protein was subsequently tested for enzymatic properties and revealed to exhibit its highest activity at pH 8 and 40 C. Using homology modeling and molecular docking techniques, the three-dimensional model of SeChi was constructed and screened for inhibitors. In two rounds of screening, twenty compounds were selected. With the purified rSeChi, we tested each of the twenty compounds for inhibitor activity against rSeChi, and seven compounds showed obvious activity. This study provided new information for the chitinase of beet armyworm and for chitinase inhibitor development.

The haemagglutinin (HA) gene of H5N1 Avian influenza virus (AIV) was amplified from the plasmid pGEM-HA and cloned into the baculovirus transfer plasmid pFastBacHT to construct the recombinant transfer vector pFastBacHT-HA. The pFastBacHT-HA was transformed into DH10Bac competent cells, transposed with a shuttle vector (Bacmid) and the transposition rBacmid-HA was constructed. The recombinant baculovirus was harvested from sf9 cells transfected with rBacmid-HA. The expressed HA protein was identified and analysed by SDS–PAGE, Western blot and haemadsorption assay. The 66 kDa protein could only react with chicken serum positive to H5 subtype AIV. The haemadsorption assay showed that the sf9 cells infected with rBacmid-HA baculovirus could absorb chicken red blood cells. These results indicated that the HA protein was successfully expressed in sf9 cells, with reaction specificity to H5 subtype antiserum.

East Coast fever (ECF) is a fatal disease of cattle caused by the protozoan parasite Theileria parva. The development of a subunit vaccine, based on the sporozoite-specific surface antigen p67, has been hampered by difficulties in achieving high-level expression of recombinant p67 in a near-authentic form. Therefore two sets of recombinant baculovirus vectors were constructed. The first set, encoding various regions of p67, produced low levels of the corresponding p67 domains in High Five™ cells, despite the presence of large amounts of p67 RNA. The second, consisting of p67 domains fused to the carboxy-terminus of GFP expressed significantly higher levels of p67 protein. The GFP[ratio ]p67 fusion proteins were recognized by a sporozoite-neutralizing monoclonal antibody (TpM12) raised against native p67 whereas non-fused full length p67 expressed in insect cells was not recognized. GFP-tagging therefore, appeared to enhance the stability of p67 and to conserve its folding. The high-level expression of p67 domains in a more authentic form is an important step towards the development of an effective subunit vaccine against ECF.