title: IQcat: multiplexed protein quantification by isoelectric QconCAT.
abstract: Expression of isotopically labeled peptide standards as artificial concatamers (QconCATs) allows for the multiplex quantification of proteins in unlabeled samples by mass spectrometry. We have developed a generalizable QconCAT design strategy, which we term IQcat, wherein concatenated peptides are binned by pI to facilitate MS-sample enrichment by isoelectric focusing. Our method utilizes a rapid (∼2 weeks), inexpensive and scalable purification of arg/lys labeled IQcat standards in the Escherichia coli auxotroph AT713. With this pipeline, we assess the fidelity of IQcat-based absolute quantification for ten yeast proteins over a broad concentration range in a single information-rich isoelectric fraction. The technique is further employed for a quantitative study of androgen-dependent protein expression in cultured prostate cancer cells.
authors.affiliation: Institute for Systems Biology, Seattle, WA, USA.
authors.forename: Ryan J
authors.initials: RJ
authors.lastname: Austin
authors.forename: Deborah K
authors.initials: DK
authors.lastname: Chang
authors.forename: Carly A
authors.initials: CA
authors.lastname: Holstein
authors.forename: Lik W
authors.initials: LW
authors.lastname: Lee
authors.forename: Jenni
authors.initials: J
authors.lastname: Risler
authors.forename: Jonathan H
authors.initials: JH
authors.lastname: Wang
authors.forename: Lee
authors.initials: L
authors.lastname: Adams
authors.forename: Nicolle B
authors.initials: NB
authors.lastname: Krusberski
authors.forename: Daniel B
authors.initials: DB
authors.lastname: Martin
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D045744:Cell Line, Tumor; D006801:Humans; D007525:Isoelectric Focusing / Q000191:economics / Q000379:methods*; D008297:Male; D013058:Mass Spectrometry / Q000191:economics / Q000379:methods*; D008969:Molecular Sequence Data; D010455:Peptides / Q000032:analysis*; D011471:Prostatic Neoplasms / Q000737:chemistry; D011506:Proteins / Q000032:analysis*; D040901:Proteomics / Q000191:economics / Q000379:methods*; D012441:Saccharomyces cerevisiae / Q000737:chemistry; D029701:Saccharomyces cerevisiae Proteins / Q000032:analysis
chemical_list: D010455:Peptides; D011506:Proteins; D029701:Saccharomyces cerevisiae Proteins