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Figure 1. Representative 2D gels of Macaca mulatta protein sample stained with SyproRuby™. The polypeptide molecular mass scale in kDa is depicted on the y-axis while the x-axis shows the pI range. The proteins were resolved in 4–7 linear pH gradient (Immobiline DryStrips; 240 × 3 × 0.5) and 8–15% gradient SDS-PAGE (2400 × 2000 × 1 mm). The results of the proteins that were identified (indicated by arrows) by in-gel trypsin digestion and MALDI-TOF-TOF followed by de novo sequencing are elaborated in Table 1.
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Figure 2. Representative De novo analysis of a MALDI-TOF-TOF spectrum. (A) The x- and y-axes show the mass to charge (m/z) ratio and the % abundance of the precursor ion fragments, respectively. The MS/MS spectrum was analyzed by PEAKS de novo sequencing software to generate 'RSALQAAHDAVAQEGQCR'. (B) The table details peptide fragments 'b-ions, y-ions and a-ions' and the neutral losses of water and ammonia for b-ions and y-ions as well as the immonium ions to develop confident and complete peptide sequence de novo from MS/MS spectrum. (C) The SPIDER homology search of this peptide resulted in sequence from Ubiquitin carboxyl-terminal hydrolase isozyme L1 as 'NEAIQAAHDAVAQEGQCR'.
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Figure 3. Schematic of the methodology for compilation of protein database for Macaca mulatta from De Novo analysis of MALDI-TOF-TOF spectra.
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