Peptide libraries for proteomics
Peptide libraries are also used for proteomics. Indeed, quantitative proteomics often consists in the absolute quantification of one or more peptides of interest in biological samples. Libraries of heavy isotope peptides are ideal for proteomics assays. These are isotopically labeled peptides, also named Stable isotope labeled peptides (SIL) or heavy peptides, in order to be able to differentiate from the peptide of interest only by a higher mass. Thus, libraries of heavy isotope peptides are used as a chemical reference substance to quantify the peptide in given biological samples.
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Bohrer BC, Li YF, Reilly JP, et al. Combinatorial libraries of synthetic peptides as a model for shotgun proteomics. Anal Chem. (2010)
BACKGROUND : A synthetic approach to model the analytical complexity of biological proteolytic digests has been developed. Combinatorial peptide libraries ranging in length between nine and twelve amino acids that represent typical tryptic digests were designed, synthesized and analyzed.
METHODS : Individual libraries and mixtures thereof were studied by replicate liquid chromatography-ion trap mass spectrometry and compared to a tryptic digest of Deinococcus radiodurans. Similar to complex proteome analysis, replicate study of individual libraries identified additional unique peptides. Fewer novel sequences were revealed with each additional analysis in a manner similar to that observed for biological data.
RESULTS : Our results demonstrate a bimodal distribution of peptides sorting to either very low or very high levels of detection. Upon mixing of libraries at equal abundance, a length-dependent bias in favor of longer sequence identification was observed. Peptide identification as a function of site-specific amino acid content was characterized with certain amino acids proving to be of considerable importance.
CONCLUSION : This report demonstrates that peptide libraries of defined character can serve as a reference for instrument characterization in proteomics studies. Furthermore, they are uniquely suited to delineate the physical properties that influence identification of peptides which provides a foundation for optimizing the study of samples with less defined heterogeneity.
Andrew N. Hoofnagle and Cory Bystrom, Proteomics, Principles and Applications of Clinical Mass Spectrometry, 10.1016/B978-0-12-816063-3.00006-2, (181-201), (2018).
BACKGROUND : This paper describes an algorithm to apply proteotypic peptide sequence libraries to protein identifications performed using tandem mass spectrometry (MS/MS). Proteotypic peptides are those peptides in a protein sequence that are most likely to be confidently observed by current MS-based proteomics methods.
METHODS : Libraries of proteotypic peptide sequences were compiled from the Global Proteome Machine Database for Homo sapiens and Saccharomyces cerevisiae model species proteomes. These libraries were used to scan through collections of tandem mass spectra to discover which proteins were represented by the data sets, followed by detailed analysis of the spectra with the full protein sequences corresponding to the discovered proteotypic peptides.
RESULTS : This algorithm (Proteotypic Peptide Profiling, or P3) resulted in sequence-to-spectrum matches comparable to those obtained by conventional protein identification algorithms using only full protein sequences, with a 20-fold reduction in the time required to perform the identification calculations. The proteotypic peptide libraries, the open source code for the implementation of the search algorithm and a website for using the software have been made freely available. Approximately 4% of the residues in the H. sapiens proteome were required in the proteotypic peptide library to successfully identify proteins.