Assessment of cardiac proteome dynamics with heavy water

Slower protein synthesis rates in interfibrillar than subsarcolemmal mitochondria

Takhar Kasumov, Erinne R. Dabkowski, Kadambari Chandra Shekar, Ling Li, Rogerio F. Ribeiro, Kenneth Walsh, Stephen F. Previs, Rovshan Sadygov, Belinda Willard, William C. Stanley

Research output: Contribution to journalArticle

39 Citations (Scopus)

Abstract

Traditional proteomics provides static assessment of protein content, but not synthetic rates. Recently, proteome dynamics with heavy water (2H2O) was introduced, where 2H labels amino acids that are incorporated into proteins, and the synthesis rate of individual proteins is calculated using mass isotopomer distribution analysis. We refine this approach with a novel algorithm and rigorous selection criteria that improve the accuracy and precision of the calculation of synthesis rates and use it to measure protein kinetics in spatially distinct cardiac mitochondrial subpopulations. Subsarcolemmal mitochondria (SSM) and interfibrillar mitochondria (IFM) were isolated from adult rats, which were given 2H2O in the drinking water for up to 60 days. Plasma 2H2O and myocardial 2H-enrichment of amino acids were stable throughout the experimental protocol. Multiple tryptic peptides were identified from 28 proteins in both SSM and IFM and showed a time-dependent increase in heavy mass isotopomers that was consistent within a given protein. Mitochondrial protein synthesis was relatively slow (average half-life of 30 days, 2.4% per day). Although the synthesis rates for individual proteins were correlated between IFM and SSM (R2 = 0.84; P < 0.0001), values in IFM were 15% less than SSM (P < 0.001). In conclusion, administration of 2H2O results in stable enrichment of the cardiac precursor amino acid pool, with the use of refined analytical and computational methods coupled with cell fractionation one can measure synthesis rates for cardiac proteins in subcellular compartments in vivo, and protein synthesis is slower in mitochondria located among the myofibrils than in the subsarcolemmal region.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Heart and Circulatory Physiology
Volume304
Issue number9
DOIs
StatePublished - 2013

Fingerprint

Deuterium Oxide
Proteome
Mitochondria
Proteins
Amino Acids
Cell Fractionation
Myofibrils
Mitochondrial Proteins
Drinking Water
Proteomics
Patient Selection
Half-Life
Peptides

Keywords

  • Deuterium
  • Heart
  • Heavy water
  • High resolution mass spectroscopy
  • Proteome dynamics
  • Proteomics

ASJC Scopus subject areas

  • Physiology
  • Physiology (medical)
  • Cardiology and Cardiovascular Medicine

Cite this

Assessment of cardiac proteome dynamics with heavy water : Slower protein synthesis rates in interfibrillar than subsarcolemmal mitochondria. / Kasumov, Takhar; Dabkowski, Erinne R.; Shekar, Kadambari Chandra; Li, Ling; Ribeiro, Rogerio F.; Walsh, Kenneth; Previs, Stephen F.; Sadygov, Rovshan; Willard, Belinda; Stanley, William C.

In: American Journal of Physiology - Heart and Circulatory Physiology, Vol. 304, No. 9, 2013.

Research output: Contribution to journalArticle

Kasumov, Takhar ; Dabkowski, Erinne R. ; Shekar, Kadambari Chandra ; Li, Ling ; Ribeiro, Rogerio F. ; Walsh, Kenneth ; Previs, Stephen F. ; Sadygov, Rovshan ; Willard, Belinda ; Stanley, William C. / Assessment of cardiac proteome dynamics with heavy water : Slower protein synthesis rates in interfibrillar than subsarcolemmal mitochondria. In: American Journal of Physiology - Heart and Circulatory Physiology. 2013 ; Vol. 304, No. 9.
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AU - Shekar, Kadambari Chandra

AU - Li, Ling

AU - Ribeiro, Rogerio F.

AU - Walsh, Kenneth

AU - Previs, Stephen F.

AU - Sadygov, Rovshan

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AU - Stanley, William C.

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AB - Traditional proteomics provides static assessment of protein content, but not synthetic rates. Recently, proteome dynamics with heavy water (2H2O) was introduced, where 2H labels amino acids that are incorporated into proteins, and the synthesis rate of individual proteins is calculated using mass isotopomer distribution analysis. We refine this approach with a novel algorithm and rigorous selection criteria that improve the accuracy and precision of the calculation of synthesis rates and use it to measure protein kinetics in spatially distinct cardiac mitochondrial subpopulations. Subsarcolemmal mitochondria (SSM) and interfibrillar mitochondria (IFM) were isolated from adult rats, which were given 2H2O in the drinking water for up to 60 days. Plasma 2H2O and myocardial 2H-enrichment of amino acids were stable throughout the experimental protocol. Multiple tryptic peptides were identified from 28 proteins in both SSM and IFM and showed a time-dependent increase in heavy mass isotopomers that was consistent within a given protein. Mitochondrial protein synthesis was relatively slow (average half-life of 30 days, 2.4% per day). Although the synthesis rates for individual proteins were correlated between IFM and SSM (R2 = 0.84; P < 0.0001), values in IFM were 15% less than SSM (P < 0.001). In conclusion, administration of 2H2O results in stable enrichment of the cardiac precursor amino acid pool, with the use of refined analytical and computational methods coupled with cell fractionation one can measure synthesis rates for cardiac proteins in subcellular compartments in vivo, and protein synthesis is slower in mitochondria located among the myofibrils than in the subsarcolemmal region.

KW - Deuterium

KW - Heart

KW - Heavy water

KW - High resolution mass spectroscopy

KW - Proteome dynamics

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