Design of signal peptide bombyxin and its effect on secretory expression efficiency and levels of Helicobacter pylori urease subunit B in silkworm cells and larvae

• Published in: Brazilian Archives of Biology and Technology Vol. 58; no. 3; pp. 319 - 325
• Main Authors: Zhang, Xiao-lin, Jiang, An-min, Ma, Zhong-you, Xiong, You-yi, Dou, Jin-feng, Zhou, Guo-liang, Qin, Mei-song, Wang, Jian-fei
• Format: Journal Article
• Language: English
• Published: Instituto de Tecnologia do Paraná (Tecpar) 01.06.2015
• Subjects: baculovirus expression system; Bombyx mori; Helicobacter pylori urease subunit B; secretory expression; signal peptide
• ISSN: 1516-8913; 1678-4324; 1516-8913
• DOI: 10.1590/S1516-8913201500013

This study employed a Bac-to-Bac/Bombyx mori bioreactor to mass-produce immunogenic urease subunit B (UreB) from Helicobacter pylori.The signal peptide bombyxin from B. mori was used to promote secretory expression to improve expression levels and was designed and integrated into the UreB gene to generate the Bacmid/BmNPV/(signal peptide)-UreB baculovirus expression system. To determine whether the bombyxin signal peptide resulted in secretory expression of recombinant UreB (rUreB) and to determine the secretory efficiency, we tested the secretory expression level of rUreB in Bm5 cells using ELISA. To further investigate whether secretory expression affected cell viability, cells were evaluated using 0.4% trypan blue staining, and Bacmid/BmNPV/UreB without the signal peptide served as a control. The above recombinant bacmid constructs were injected to silkworm larvae, and the secretory expression level of rUreB was detected using SDS-PAGE and semi-quantitative western blot analysis. The results indicated that the bombyxin signal peptide directed the secretory expression of rUreB and that this expression improved the viability of Bm5 cells. Moreover, the results showed that the expression level of rUreB was 1.5 times higher with the Bacmid/BmNPV constructs containing the bombyxin signal sequence than those without the signal sequence. These results demonstrate that secretory expression can enhance rUreB expression levels and is likely to aid in the large-scale expression and yield of rUreB in silkworm larvae.