Measurement of Fractional Synthetic Rates of Multiple Protein Analytes by Triple Quadrupole Mass Spectrometry 

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dc.contributor.authorLee, Anita Y H
dc.contributor.authorYates, Nathan A
dc.contributor.authorIchetovkin, Marina
dc.contributor.authorDeyanova, Ekaterina
dc.contributor.authorSouthwick, Katie
dc.contributor.authorFisher, Timothy S
dc.contributor.authorWang, Weixun
dc.contributor.authorLoderstedt, James
dc.contributor.authorWalker, Nykia
dc.contributor.authorZhou, Haihong
dc.contributor.authorZhao, Xuemei
dc.contributor.authorSparrow, Carl P
dc.contributor.authorHubbard, Brian K
dc.contributor.authorRader, Daniel J
dc.contributor.authorSitlani, Ayesha
dc.contributor.authorMillar, John S
dc.contributor.authorHendrickson, Ronald C
dc.date.accessioned2022-03-24T17:42:58Z
dc.date.available2022-03-24T17:42:58Z
dc.date.issued2012-03-01
dc.description.abstractBACKGROUND Current approaches to measure protein turnover that use stable isotope-labeled tracers via GC-MS are limited to a small number of relatively abundant proteins. We developed a multiplexed liquid chromatography–selected reaction monitoring mass spectrometry (LC-SRM) assay to measure protein turnover and compared the fractional synthetic rates (FSRs) for 2 proteins, VLDL apolipoprotein B100 (VLDL apoB100) and HDL apoA-I, measured by both methods. We applied this technique to other proteins for which kinetics are not readily measured with GC-MS. METHODS Subjects were given a primed-constant infusion of [5,5,5-D₃]-leucine (D₃-leucine) for 15 h with blood samples collected at selected time points. Apolipoproteins isolated by SDS-PAGE from lipoprotein fractions were analyzed by GC-MS or an LC-SRM assay designed to measure the M+3/M+0 ratio at >1% D₃-leucine incorporation. We calculated the FSR for each apolipoprotein by curve fitting the tracer incorporation data from each subject. RESULTS The LC-SRM method was linear over the range of tracer enrichment values tested and highly correlated with GC-MS (R² > 0.9). The FSRs determined from both methods were similar for HDL apoA-I and VLDL apoB100. We were able to apply the LC-SRM approach to determine the tracer enrichment of multiple proteins from a single sample as well as proteins isolated from plasma after immunoprecipitation. CONCLUSIONS The LC-SRM method provides a new technique for measuring the enrichment of proteins labeled with stable isotopes. LC-SRM is amenable to a multiplexed format to provide a relatively rapid and inexpensive means to measure turnover of multiple proteins simultaneously.en_US
dc.description.sponsorshipWe thank James P. Conway for his expert assistance with the protein databases (merck.protein). Research Funding: Merck Sharp & Dohme Corp.en_US
dc.description.urihttps://academic.oup.com/clinchem/article/58/3/619/5620621?login=trueen_US
dc.format.extent9 pagesen_US
dc.genrejournal articlesen_US
dc.identifierdoi:10.13016/m2s6jj-xbrz
dc.identifier.citationAnita Y H Lee, Nathan A Yates, Marina Ichetovkin, Ekaterina Deyanova, Katie Southwick, Timothy S Fisher, Weixun Wang, James Loderstedt, Nykia Walker, Haihong Zhou, Xuemei Zhao, Carl P Sparrow, Brian K Hubbard, Daniel J Rader, Ayesha Sitlani, John S Millar, Ronald C Hendrickson, Measurement of Fractional Synthetic Rates of Multiple Protein Analytes by Triple Quadrupole Mass Spectrometry, Clinical Chemistry, Volume 58, Issue 3, 1 March 2012, Pages 619–627, https://doi.org/10.1373/clinchem.2011.172429en_US
dc.identifier.urihttps://doi.org/10.1373/clinchem.2011.172429
dc.identifier.urihttp://hdl.handle.net/11603/24419
dc.language.isoen_USen_US
dc.publisherOxford University Pressen_US
dc.relation.isAvailableAtThe University of Maryland, Baltimore County (UMBC)
dc.relation.ispartofUMBC Biological Sciences Department Collection
dc.rightsThis item is likely protected under Title 17 of the U.S. Copyright Law. Unless on a Creative Commons license, for uses protected by Copyright Law, contact the copyright holder or the author.en_US
dc.subjectmeasuring protein turnoveren_US
dc.subjectmultiplexed liquid chromatography–selected reaction monitoring mass spectrometry (LC-SRM) assayen_US
dc.subjectfractional synthetic rates (FSRs)en_US
dc.titleMeasurement of Fractional Synthetic Rates of Multiple Protein Analytes by Triple Quadrupole Mass Spectrometry en_US
dc.typeTexten_US
dcterms.creatorhttps://orcid.org/0000-0001-7603-9276en_US

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