Expression of Trichoderma viride endoglucanase III in the larvae of silkworm, Bombyx mori L. and characteristic analysis of the recombinant protein

• DOI: 10.1007/s11033-010-0505-4
• Publication Date: 2010-11-24T08:17:36Z
• Authors: Xing-hua Li; Peng Zhang; Mei-xian Wang; Fang Zhou; Firdose Ahmad Malik; Hua-jun Yang; Roy Bhaskar; Jia-biao Hu; Chun-guang Sun; Yun-gen Miao
• Keywords: Trichoderma; 0303 health sciences; Blotting, Western; Gene Expression; Exons; Bombyx; Polymerase Chain Reaction; Introns; Recombinant Proteins; Cell Line; 03 medical and health sciences; Cellulase; Larva; Animals; Baculoviridae

Endoglucanase is a part of cellulase which hydrolyzes cellulose into glucose. In this study, we cloned endoglucanase III (EG III) gene from Trichoderma viride strain AS 3.3711 using a PCR-based exon splicing method, and expressed EG III recombinant protein in both silkworm BmN cell line and silkworm larvae with an improved Bac-to-Bac/BmNPV mutant baculovirus expression system, which lacks the chiA and v-cath genes of Bombyx mori nucleopolyhedrovirus (BmNPV). The result showed that around 45 kDa protein was visualized in BmN cells at 48 h after the second generation recombinant mBacmid/BmNPV/EG III baculovirus infection. The enzymes from recombinant baculoviruses infected silkworms exhibited significant maximum enzyme activity at the environmental condition of pH 8.0 and temperature 50°C, and increased 20.94 and 19.13% compared with that from blank mBacmid/BmNPV baculoviruses infected silkworms and normal silkworms, respectively. It was stable at pH range from 5.0 to 9.0 and at temperature range from 40 to 60°C. It provided a possibility to generate transgenic silkworms expressing bio-active cellulase, which can catabolize dietary fibers more efficiently, and it might be of great significance for sericulture industry.