Multiple apolipoprotein kinetics measured in human HDL by high-resolution/accurate mass parallel reaction monitoring[S]

• Published in: Journal of lipid research Vol. 57; no. 4; pp. 714 - 728
• Main Authors: Singh, Sasha A., Andraski, Allison B., Pieper, Brett, Goh, Wilson, Mendivil, Carlos O., Sacks, Frank M., Aikawa, Masanori
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01-04-2016; The American Society for Biochemistry and Molecular Biology; Elsevier
• Subjects: Adult; Amino Acid Sequence; Cardiovascular Diseases - metabolism; compartmental modeling; coronary heart disease; Female; Humans; Kinetics; Lipoproteins, HDL - chemistry; Lipoproteins, HDL - metabolism; Male; Mass Spectrometry; metabolism; Methods; Middle Aged; Particle Size; Peptide Fragments - chemistry; Peptide Fragments - metabolism; Proteomics - methods; Risk Factors; size fractionation; size fractionation; compartmental modeling; mass spectrometry; coronary heart disease; metabolism
• ISSN: 0022-2275; 1539-7262
• DOI: 10.1194/jlr.D061432

Endogenous labeling with stable isotopes is used to study the metabolism of proteins in vivo. However, traditional detection methods such as GC/MS cannot measure tracer enrichment in multiple proteins simultaneously, and multiple reaction monitoring MS cannot measure precisely the low tracer enrichment in slowly turning-over proteins as in HDL. We exploited the versatility of the high-resolution/accurate mass (HR/AM) quadrupole Orbitrap for proteomic analysis of five HDL sizes. We identified 58 proteins in HDL that were shared among three humans and that were organized into five subproteomes according to HDL size. For seven of these proteins, apoA-I, apoA-II, apoA-IV, apoC-III, apoD, apoE, and apoM, we performed parallel reaction monitoring (PRM) to measure trideuterated leucine tracer enrichment between 0.03 to 1.0% in vivo, as required to study their metabolism. The results were suitable for multicompartmental modeling in all except apoD. These apolipoproteins in each HDL size mainly originated directly from the source compartment, presumably the liver and intestine. Flux of apolipoproteins from smaller to larger HDL or the reverse contributed only slightly to apolipoprotein metabolism. These novel findings on HDL apolipoprotein metabolism demonstrate the analytical breadth and scope of the HR/AM-PRM technology to perform metabolic research.