Static and turnover kinetic measurement of protein biomarkers involved in triglyceride metabolism including apoB48 and apoA5 by LC/MS/MS.
LC/MS quantification of multiple plasma proteins that differ by several orders of magnitude in concentration from a single sample is challenging. We present a strategy that allows the simultaneous determination of the concentration and turnover kinetics of higher and lower abundant proteins from a single digestion mixture. Our attention was directed at a cluster of proteins that interact to affect the absorption and interorgan lipid trafficking. We demonstrate that apos involved in TG metabolism such as apoC2, C3, E, and A4 (micromolar concentration), and apoB48 and apoA5 (single-digit nanomolar concentration) can be quantified from a single digestion mixture. A high degree of correlation between LC/MS and immunobased measurements for apoC2, C3, E, and B48 was observed. Moreover, apoA5 fractional synthesis rate was measured in humans for the first time. Finally, the method can be directly applied to studies involving nonhuman primates because peptide sequences used in the method are conserved between humans and nonhuman primates.
Merck Research Laboratories, Merck & Co. Inc., Kenilworth, NJ.
Yi
Y
Pan
Merck Research Laboratories, Merck & Co. Inc., Kenilworth, NJ.
Haihong
H
Zhou
Merck Research Laboratories, Merck & Co. Inc., Kenilworth, NJ.
Ablatt
A
Mahsut
Merck Research Laboratories, Merck & Co. Inc., Kenilworth, NJ.
Rory J
RJ
Rohm
Merck Research Laboratories, Merck & Co. Inc., Kenilworth, NJ.
Olga
O
Berejnaia
Merck Research Laboratories, Merck & Co. Inc., Kenilworth, NJ.
Olga
O
Price
Merck Research Laboratories, Merck & Co. Inc., Kenilworth, NJ.
Ying
Y
Chen
Merck Research Laboratories, Merck & Co. Inc., Kenilworth, NJ.
Jose
J
Castro-Perez
Merck Research Laboratories, Merck & Co. Inc., Kenilworth, NJ.
Michael E
ME
Lassman
Merck Research Laboratories, Merck & Co. Inc., Kenilworth, NJ.
David
D
McLaren
Merck Research Laboratories, Merck & Co. Inc., Kenilworth, NJ.
James
J
Conway
Merck Research Laboratories, Merck & Co. Inc., Kenilworth, NJ.
Kristian K
KK
Jensen
Columbia University, New York, NY.
Tiffany
T
Thomas
Columbia University, New York, NY.
Gissette
G
Reyes-Soffer
Columbia University, New York, NY.
Henry N
HN
Ginsberg
Merck Research Laboratories, Merck & Co. Inc., Kenilworth, NJ.
David E
DE
Gutstein
Merck Research Laboratories, Merck & Co. Inc., Kenilworth, NJ.
Michele
M
Cleary
Merck Research Laboratories, Merck & Co. Inc., Kenilworth, NJ.
Stephen F
SF
Previs
Merck Research Laboratories, Merck & Co. Inc., Kenilworth, NJ.
Thomas P
TP
Roddy
D000072040:Apolipoprotein A-V / Q000097:blood*; D053283:Apolipoprotein B-48 / Q000097:blood*; D015415:Biomarkers / Q000097:blood; D002853:Chromatography, Liquid / Q000379:methods; D005260:Female; D006801:Humans; D008297:Male; D013058:Mass Spectrometry / Q000379:methods; D014280:Triglycerides / Q000097:blood*
C437704:APOA5 protein, human; D000072040:Apolipoprotein A-V; D053283:Apolipoprotein B-48; D015415:Biomarkers; D014280:Triglycerides
dyslipidemia; kinetic biomarker; mass spectrometry; stable isotopes