DOI: 10.1101/472662Nov 19, 2018Paper

Screening a Resource of Recombinant Protein Fragments for Targeted Proteomics

BioRxiv : the Preprint Server for Biology
Fredrik EdforsMathias Uhlén

Abstract

The availability of proteomics resources hosting protein and peptide standards, as well as the data describing their analytical performances, will continue to enhance our current capabilities to develop targeted proteomics methods for quantitative biology. This study describes the analysis of a resource of 26,840 individually purified recombinant protein fragments corresponding to more than 16,000 human protein-coding genes. The resource was screened to identify proteotypic peptides suitable for targeted proteomics efforts and we report LC-MS/MS assay coordinates for more than 25,000 proteotypic peptides, corresponding to more than 10,000 unique proteins. Additionally, peptide formation and digestion kinetics were, for a subset of the standards, monitored using a time-course protocol involving parallel digestion of isotope-labelled recombinant protein standards and endogenous human plasma proteins. We show that the strategy by adding isotope-labelled recombinant proteins prior to trypsin digestion enables short digestion protocols (≤60 min) with robust quantitative precision. In a proof-of-concept study, we quantified 23 proteins in human plasma using assay parameters defined in our study and used the standards to describe dist...Continue Reading

Related Concepts

Digestion
Isotope Labeling
Peptides
Plasma
Recombinant Proteins
Trypsin
LPAR4 protein, human
Proteomics
Liquid Chromatography Mass Spectrometry
Analysis

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