Neuropeptide identification in mass spectrometry experiments using database search programs developed for proteins is challenging. Unlike proteins, the detection of the complete sequence using a single spectrum is required to identify neuropeptides or prohormone peptides. This study compared the performance of three open-source programs used to identify proteins, OMSSA, X!Tandem and Crux, to identify prohormone peptides. From a target database of 7850 prohormone peptides, 23550 query spectra were simulated across different scenarios. Crux was the only program that correctly matched all peptides regardless of p-value and at p-value < 1 × 10-2, 33%, 64%, and >75%, of the 5, 6, and ≥7 amino acid-peptides were detected. Crux also had the best performance in the identification of peptides from chimera spectra and in a variety of missing ion scenarios. OMSSA, X!Tandem and Crux correctly detected 98.9% (99.9%), 93.9% (97.4%) and 88.7% (98.3%) of the peptides at E- or p-value < 1 × 10-6 (< 1 × 10-2), respectively. OMSSA and X!Tandem outperformed the other programs in significance level and computational speed, respectively. A consensus approach is not recommended because some prohormone peptides were only identified by one program.
- database search program
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