Identification and Molecular Characterization of Inducible Immune Protein in Bombyx mori L

Introduction: In insect immunity, antibacterial proteins are an important part of the immune system. These proteins are mostly produced by epithelial cells and released through hemolymph. Antibacterial proteins in insects belong to the five major families of cecropins, defensins, attacinlike proteins, proline-rich peptides, and lysozymes. Considering the importance of these proteins in fighting infections, the aim of this study was to evaluate and identify these proteins in silkworms (Bombyx mori).Materials and Methods: In the present work, profiling of the proteins present in the hemolymph of control silkworms versus, those infected with bacteria was performed by SDS-PAGE, ۲D gel electrophoresis, and image analysis. We also used MALDI-TOF and MS/MS to investigate novel and uncharacterized immune protein. For this aim, the silkworm hemolymph after inoculation and infection with Staphylococcus aureus bacteria was analyzed using SDS-PAGE, ۲-dimensional gel electrophoresis, MALDI-TOF and MS/MS to identify the immune proteins. The Swiss-Prot and NCBI databases were used for protein identification.Results: A novel protein with a molecular weight of ۳۱.۹ KDa was discovered on the fourth day of exposure to bacteria. The expressed protein showed effective activity against S. aureus, which was infected silkworms. In the MALDI-TOF/MS result and protein identification analysis, ۹۰ numbers of mass values were searched, the mass values matched ۱۶, the total sequence coverage was ۴۶%, and their score was ۵۷. According to the analyses, the expressed protein belongs to the hemolysin secretion protein (HlyD).Conclusions: The results led to the identification of a new protein with antimicrobial properties in silkworm, although more information is needed.

Keywords:

Antibacterial Proteins , Immunization , Electrophoresis , MALDI-TOF , MS-MS