Abstract: Insect carboxylesterases are important enzymes involved in xenobiotic metabolism and degradation of odorants. In the present study, Bmae35 gene, which is expressed in larval olfactory tissues of the silkworm and the putative ortholog of antennal esterase Snon-EST in Sesamia nonagrioides, was cloned and exogenously expressed. Bmae35 comprises a complete coding sequence of 1 581 bp and codes 526 amino acids. The multiple sequence alignment of Bmae35 with other esterases revealed that Bmae35 has the structure characteristics of insect esterases, including the catalytic triad (Ser191, Glu313 and His429) and the conserved pentapeptide Gly-x-Ser-x-Gly in the α-esterase family. Based on the semi-quantitative RT-PCR analysis, the expression pattern of Bmae35 gene showed that it was highly expressed in head, fat body, Malpighian tubules, integument, and silk glands in day-3 5th instar larvae. In addition, the expression of Bmae35 showed a positive correlation with sex pheromone content. The results suggest that Bmae35 might play important roles in pheromone synthesis. The Bmae35 gene was sub-cloned into the expression vector pET28(a) and the recombinant protein was obtained by isopropyl β-D-1-thiogalactopyranoside (IPTG) induction. Electrophoresis analysis showed that Bmae35 was expressed in Escherichia coli as inclusion body. The recombinant protein Bmae35 was purified by immobilized Ni²⁺ absorption chromatograph column. Western blotting analysis indicated that Bmae35 was correctly expressed in E. coli and purified. The results of this study provide useful data for further understanding the odorant detoxification and expression orientation.

Key words: Bombyx mori, carboxylesterase, gene cloning, expression pattern, prokaryotic expression, Western blotting