Abstract. Mass spectrometry (MS)-based shotgun proteomics allows protein identifications even in complex biological samples. Protein abundances can then be estimated from the counts of MS/MS spectra attributable to each protein, provided one accounts for differential MS-detectability of contributing peptides. We developed a method, APEX, which calculates Absolute Protein EXpression levels based upon learned correction factors, MS/MS spectral counts, and each protein's probability of correct identification.
The APEX protocol describes APEX-based calculations in three parts:
1. Using training data, peptide sequences and their sequence properties, a model is built to estimate MS-detectability (Oi) for any given protein.
2. Absolute protein abundances are calculated from spectral counts, identification probabilities and the learned Oi-values.
3. Simple statistics allow calculation of differential expression in two distinct biological samples, i.e. measuring relative protein abundances.
APEX-based protein abundances span 3-4 orders of magnitude and are applicable to mixtures of 100s to 1000s of proteins.
Protocol paper (Nature Protocols, 2008).
Link to journal website.
Supplementary Notes (to Protocol)
Original citation: Lu, Vogel, Wang, Yao, Marcotte. Absolute protein expression profiling estimates the relative contributions of transcriptional and translational regulation. Nat Biotechnol. 2007 Jan;25(1):117-24
This website provides scripts and data files to conduct analyses described in the APEX Protocol.
Note (August 29, 2008): I am actively working on different ways to calculate significance of differential protein expression. There will be an updated perl script and a more detailed explanaition soon. C.V.Example: yeast grown in YMD and YPD (analyzed on LCQ) - data from OPD(opd00038_YEAST...opd00042_YEAST and opd00047_YEAST...opd00098_YEAST)