Proteomic analysis of silkworm fat body

Proteomic analysis was performed on the fat body of the fifth instar female silkworm (Bombyx mori, p50) larvae because it is thought that numerous biological substances are synthesized in the fat body. The Drosophila genome and silkworm expression sequence tag (EST) data were applied to the analysis of the fat body proteins and served as the database for protein identification. Protein spots were cut manually from two-dimensional polyacrylamide gel electrophoresis gels, and these spots were treated with 4-vinylpyridine for alkylation. Each spot was analyzed by capillary HPLC coupled with ion-trap mass spectrometry (MS) after proteolysis using modified trypsin. A total of 177 proteins analyzed by MS were identified using the Drosophila genome and silkworm EST data, and these proteins were mapped on 2D-gel. The fat metabolism-related proteins identified in this study were diacylglycerol binding protein, triacylglycerol lipase, and putative hydrolases. Fewer proteins than expected were identified by fat body proteomic analysis. However, nine glycolysis-related proteins were identified in the extracts of the fat bodies. Many cytoskeleton proteins and defense-related proteins were observed in the protein profile. However, some of these proteins were thought to have been derived from the muscles on the epidermis. Calreticulins and chaperonins, including heat-shock proteins, were observed in the fat body extracts.