MS identification of the Cry proteins in Bacillus thuringiensis CT-43

In Bacillus thuringiensis CT-43, five insecticidal crystal proteins (ICPs, Cry protein) are encoded. We extracted the Cry proteins, ran the SDS PAGE (two Cry protein bands were observed), and tried to identify the composition of the two Cry protein bands in the SDS PAGE. The bioinformatics pipeline is described as follows: First, we converted the original mass spectrum files to the mgf file (peaks file), then the mgf files were searched against the Bacillusthuringiensis CT-43 protein database using Mascot (version 2.3.02). The search parameters were: i) trypsin was chosen as the enzyme with one missed cleavage allowed; ii) the fixed modifications of carbamidomethylation were set as Cys, and variable modifications of oxidation as Met; iii) peptide tolerance was set as 0.05 Da, and MS/MS tolerance was set as 0.1 Da. The peptide charge was set as Mr, and monoisotopic mass was chosen. An automatic decoy database search strategy was employed to estimate the false discovery rate (FDR). The FDR was calculated as the false positive matches divided by the total matches. In the final search results, the FDR was less than 1.5%.