This figure displays the phenetic #click here randurls[1|1|,|CHEM1|]# grouping of: (A) the seven serotypes, most common strains, toxin sequences; (B) individually compared toxin domains of/G and the/B2 Prevot strain, the toxin sequence in the/B family that shares the most similarities with/G; (C) the seven serotypes,

most common strains, NTNH sequences; (D) the seven serotypes, most common strains, HA70 sequences; and (E) the seven serotypes, most common strains, HA17 sequences. Of the seven serotypes,/G shares the most similarity with the/B serotype. The percent identity shared between each/G and/B protein or domain is highlighted above1. Gel LC-MS/MS Analysis identified the four main proteins within the BoNT complex Six of the 17 gel GSK3326595 clinical trial slices, tryptically digested overnight and analyzed by use of nLC-MS/MS, returned protein matches with high sequence coverage and a 99% identity confidence when searched by use of PLGS v2.3 and validated with Scaffold v2.1. The four main proteins

associated with the botulinum neurotoxin complex were identified in various bands from the gel: BoNT/G, NTNH, HA70, and HA17 (Figure 4). Figure 4 1D SDS-PAGE and in gel digestion analysis of/G complex. This image depicts the All Blue standard (Bio-Rad, CA) and the/G complex after staining with GelCode™ Blue Safe Protein Stain (Pierce, IL). The lane of interest was cut into 17 segments, digested overnight, analyzed on a nanoLC-MS/MS system,

and identified by use of PLGS protein database searching. The proteins identified were BoNT/G (band 4), NTNH (5); HA70 was identified in three bands (7, 9, and 13) and HA17 in band 14. In solution Tryptic Digestion Analysis improved protein sequence coverage The results of the six digests of BoNT/G from both analytical instruments (QTof-Premier and LTQ-Orbitrap) were compiled to determine the greatest percent of sequence coverage Clomifene of each protein identified: BoNT/G [NCBI, CAA52275], NTNH [NCBI, CAA61228], HA70 [NCBI, CAA61225], and HA17 [NCBI, CAA61226] (Figure 5A-D). The percent recovery was determined by combining all unique peptides identified by both nLC-MS/MS instruments and calculating the ratio of amino acids identified vs. total amino acids in the protein sequence. Figure 5 Sequence coverage returned from in solution tryptic digests. The four main proteins that are associated with the BoNT/G complex and the percent of each sequence that was returned after digestion are highlighted above. The percent recovery was determined by combining all unique peptides returned from two nanoLC-MS/MS instruments and calculated by use of the number of amino acids recovered vs. total amino acids in the protein sequence.