Fig. 2

Mapping and sequencing GluC protease resistant fragments of R2Bm protein. Symbols and abbreviations are as in Fig. 1. a R2Bm protein was digested with GluC protease and analyzed by SDS-PAGE. Major observed bands were designated GA-GK. b Identification of proteolytic fragments of R2Bm protein. Bands from panel A were cut out, further processed, and analyzed by nano-LC-ESI-MS/MS sequencing. The N-terminal of the band producing fragment was identified by acetylation. c Map of the band purified R2Bm fragments. The major GluC generated R2Bm fragments detected in panel A are mapped below the ORF diagram and rulers. d Heatmap of GluC cleavages found in non-fractionated digestion reactions of R2Bm protein across time. Each column of boxes represents a GluC cleavage site. GluC cleaves after an E residue, indicated by a dot above the column, or a D residue (no dot). The positions of the amino terminal ends generated by observable GluC cleavages are given below the boxes. The number of peptide spectral matches (# PSM) are color coded as shown in the key. The R2Bm ORF is diagramed below the heatmap. Each row is a different time point, with the top row being the zero time point (no GluC) and the bottom row being eight hours. The triangle represents increasing time of GluC digestion