title: Quantitative, time-resolved proteomic analysis by combining bioorthogonal noncanonical amino acid tagging and pulsed stable isotope labeling by amino acids in cell culture.
abstract: An approach to proteomic analysis that combines bioorthogonal noncanonical amino acid tagging (BONCAT) and pulsed stable isotope labeling with amino acids in cell culture (pSILAC) provides accurate quantitative information about rates of cellular protein synthesis on time scales of minutes. The method is capable of quantifying 1400 proteins produced by HeLa cells during a 30 min interval, a time scale that is inaccessible to isotope labeling techniques alone. Potential artifacts in protein quantification can be reduced to insignificant levels by limiting the extent of noncanonical amino acid tagging. We find no evidence for artifacts in protein identification in experiments that combine the BONCAT and pSILAC methods.
authors.affiliation: Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, California;
authors.forename: John D
authors.initials: JD
authors.lastname: Bagert
authors.forename: Yushu J
authors.initials: YJ
authors.lastname: Xie
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Sweredoski
authors.forename: Yutao
authors.initials: Y
authors.lastname: Qi
authors.forename: Sonja
authors.initials: S
authors.lastname: Hess
authors.forename: Erin M
authors.initials: EM
authors.lastname: Schuman
authors.forename: David A
authors.initials: DA
authors.lastname: Tirrell
mesh_terms: D000596:Amino Acids / Q000737:chemistry* / Q000494:pharmacology; D018929:Cell Culture Techniques; D002853:Chromatography, Liquid; D006367:HeLa Cells; D006801:Humans; D007553:Isotope Labeling / Q000379:methods*; D013058:Mass Spectrometry; D010455:Peptides / Q000032:analysis; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*
chemical_list: D000596:Amino Acids; D010455:Peptides; D011506:Proteins