QconCAT

What is a QconCAT?

A QconCAT (Quantification conCATemer) is an artificial protein, generated by concatenation of proteotypic peptides.

QconCATs internal reference standards facilitate quantification at high accuracy, high sensitivity and allow quantification of low abundant proteins over a very large dynamic range.

Furthermore, by being produced as a single protein, the proteotypic peptides are always released in a ratio of 1:1 and are protected from degradation and adsorption to vessel walls.

One QconCAT can comprise of up to 100 quantotypic peptides, thus being much more cost-efficient than heavy-isotope labelled full proteins.

QconCATs can be easily integrated in established workflows and deliver highly reproducible data.

What are the applications?

QconCATs are valuable tools for various research applications involving quantitative proteomics such as: biomarker discovery and validation, systems biology, clinical research, drug discovery, bioengineering, quality control, proteomics research (e. g. regulatory networks, signalling pathways, metabolomics) etc. QconCATs are especially suited for stoichiometry analyses of pathways and protein complexes.

Advantages

  • Accurate quantification
  • Easy multiplexing (quantify a complete proteome using multiple QconCATs)
  • Scalable to 1000’s of samples/data points
  • Reproducible
  • Robust synthesis of large amounts
  • Easy integration into established quantitative protein analysis setup
  • Compatible with other methods
  • Internal sample preparation control

Selected references

Most recent publications employing QconCAT technology
  • Rimbert A, Dalila N, Wolters JC, Huijkman N, Smit M, Kloosterhuis N, Riemsma M, van der Veen Y, Singla A, van Dijk F; Biobank-Based Integrative Omics Studies Consortium, Frikke-Schmidt R, Burstein E, Tybjærg-Hansen A, van de Sluis B, Kuivenhoven JA. A common variant in CCDC93 protects against myocardial infarction and cardiovascular mortality by regulating endosomal trafficking of low-density lipoprotein receptor.
    J Eur Heart J. 2019 Oct 19 [Pubmed]
  • de Boer JF, Verkade E, Mulder NL, de Vries HD, Huijkman NC, Koehorst M, Boer T, Wolters JC, Bloks VW, van de Sluis B, Kuipers F.A. Human-like Bile Acid Pool Induced by Deletion of Cyp2c70 Modulates Effects of Farnesoid X Receptor Activation in Mice.
    J Lipid Res. 2019 Sep 10 [Pubmed]
  • Wijers M, Zanoni P, Liv N, Vos DY, Jäckstein MY, Smit M, Wilbrink S, Wolters JC, van der Veen YT, Huijkman N, Dekker D, Kloosterhuis N, van Dijk TH, Billadeau DD, Kuipers F, Klumperman J, von Eckardstein A, Kuivenhoven JA, van de Sluis B. The hepatic WASH complex is required for efficient plasma LDL and HDL cholesterol clearance.
    JCI Insight. 2019 Jun 6;4(11) [Pubmed]
  • Hoogerland JA, Lei Y, Wolters JC, de Boer JF, Bos T, Bleeker A, Mulder NL, van Dijk TH, Kuivenhoven JA, Rajas F, Mithieux G, Haeusler RA, Verkade HJ, Bloks VW, Kuipers F, Oosterveer MH. Glucose-6-Phosphate Regulates Hepatic Bile Acid Synthesis in Mice.
    Hepatology. 2019 May 18 [Pubmed]
  • Fedoseienko A, Wijers M, Wolters JC, Dekker D, Smit M, Huijkman N, Kloosterhuis N, Klug H, Schepers A, Willems van Dijk K, Levels JH, Billadeau DD, Hofker MH, van Deursen J, Westerterp M, Burstein E, Kuivenhoven JA, van de Sluis B. COMMD Family Regulates Plasma LDL Levels and Attenuates Atherosclerosis Through Stabilizing the CCC Complex in Endosomal LDLR Trafficking.
    Circ Res. 2018 Mar 15 [Pubmed]
  • Van Erp AC, Rebolledo RA, Hoeksma D, Jespersen NR, Ottens PJ, Nørregaard R, Pedersen M, Laustsen C, Burgerhof JGM, Wolters JC, Ciapaite J, Bøtker HE, Leuvenink HGD, Jespersen B. Organ-specific responses during brain death: increased aerobic metabolism in the liver and anaerobic metabolism with decreased perfusion in the kidneys.
    Sci Rep. 2018 Mar 13;8(1):4405. [Pubmed]
  • Stelder SK, Benito de Moya C, Hoefsloot HCJ, de Koning LJ, Brul S, de Koster CG. Stoichiometry, Absolute Abundance, and Localization of Proteins in the Bacillus cereus Spore Coat Insoluble Fraction Determined Using a QconCAT Approach.
    J Proteome Res. 2018 Feb 2;17(2):903-917 [Pubmed]
  • de Boer JF, Schonewille M, Boesjes M, Wolters H, Bloks VW, Bos T, van Dijk TH, Jurdzinski A, Boverhof R, Wolters JC, Kuivenhoven JA, van Deursen JM, Oude Elferink RPJ, Moschetta A, Kremoser C, Verkade HJ, Kuipers F, Groen AK. Intestinal Farnesoid X Receptor Controls Transintestinal Cholesterol Excretion in Mice.
    Gastroenterology. 2017 Apr;152(5):1126-1138 [Pubmed]
  • van Eunen K, Volker-Touw CM, Gerding A, Bleeker A, Wolters JC, van Rijt WJ, Martines AM, Niezen-Koning KE, Heiner RM, Permentier H, Groen AK, Reijngoud DJ, Derks TG, Bakker BM. Living on the edge: substrate competition explains loss of robustness in mitochondrial fatty-acid oxidation disorders.
    BMC Biol. 2016 Dec 7;14(1):107. [Pubmed]
  • Smith DG, Gingras G, Aubin Y, Cyr TD. Design and expression of a QconCAT protein to validate Hi3 protein quantification of influenza vaccine antigens.
    J Proteomics. 2016 Sep 2 [Pubmed]
  • Wolters JC, Ciapaite J, van Eunen K, Niezen-Koning KE, Matton A, Porte RJ, Horvatovich P, Bakker BM, Bischoff R, Permentier HP. Translational Targeted Proteomics Profiling of Mitochondrial Energy Metabolic Pathways in Mouse and Human Samples. J. Proteome Res. 2016 Aug 5 [Pubmed]
  • Schonewille M, de Boer JF, Mele L, Wolters H, Bloks VW, Wolters JC, Kuivenhoven JA, Tietge UJ, Brufau G, Groen AK. Statins increase hepatic cholesterol synthesis and stimulate fecal cholesterol elimination in mice.
    J Lipid Res. 2016 Aug;57(8):1455-64. [Pubmed]
  • van Zutphen T, Ciapaite J, Bloks VW, Ackereley C, Gerding A, Jurdzinski A, de Moraes RA, Zhang L, Wolters JC, Bischoff R, Wanders RJ, Houten SM, Bronte-Tinkew D, Shatseva T, Lewis GF, Groen AK, Reijngoud DJ, Bakker BM, Jonker JW, Kim PK, Bandsma RH. Malnutrition-associated liver steatosis and ATP depletion is caused by peroxisomal and mitochondrial dysfunction.
    J Hepatol. 2016 Dec;65(6):1198-1208. [Pubmed]
  • Pertl-Obermeyer H, Trentmann O, Duscha K, Neuhaus HE, Schulze WX. Quantitation of Vacuolar Sugar Transporter Abundance Changes Using QconCAT Synthtetic Peptides. Front Plant Sci. 2016 Apr 12 [Pubmed]
  • Lawless C, Holman SW, Brownridge P, Lanthaler K, Harman VM, Watkins R, Hammond DE, Miller RL, Sims PF, Grant CM, Eyers CE, Beynon RJ, Hubbard SJ. Direct and Absolute Quantification of over 1800 Yeast Proteins via Selected Reaction Monitoring. Mol Cell Proteomics. 2016 Jan 10. [Pubmed]
  • Achour B, Al-Majdoub ZB, Al Feteisi H, Elmorsi Y, Rostami-Hodjegan A, Barber J, Ten Years of QconCATs: Application of Multiplexed Quantification to Small Medically-Relevant Proteomes. International Journal of Mass Spectrometry. 2015 Nov 30; 391:93-104. [PubMed]
  • Lancrajan I, Schneider-Stock R, Naschberger E, Schellerer VS, Stürzl M, Enz R. J., Absolute quantification of DcR3 and GDF15 from human serum by LC-ESI MS. Cell Mol Med. 2015 Jul;19(7):1656-71. [PubMed]

 

Examples for QconCAT application
  1. Stelder S, Benito de Moya C, Hoefsloot HCJ, Koning LJ, Brul S, de Koster CG. Stoichiometry, Absolute Abundance, and Localization of Proteins in the Bacillus cereus Spore Coat Insoluble Fraction Determined Using a QconCAT Approach. J. Proteome Res. 2018, 17 (2), pp 903–917[Journal link]
  2. Beynon R. J., Doherty M. K., Pratt J. M., Gaskell S. J., Multiplexed absolute quantification in proteomics using artificial QCAT proteins of concatenated signature peptides. Nature Methods Vol. 2 (8), August 2005, 587-589. [download]
  3. Pratt J. M., Simpson D. M., Doherty M. K., Rivers J., Gaskell S. J., Beynon R. J., Multiplexed absolute quantification for proteomics using concatenated signature peptides encoded by QconCAT genes. Nature Protocols Vol. 1 (2), 2006, 1029-1043. [download]
  4. Al-Majdoub ZM, Carroll KM, Gaskell SJ, Barber J. Quantification of the proteins of the bacterial ribosome using QconCAT technology. J Proteome Res. 2014 Mar 7;13(3):1211-22. [PubMed]
  5. Johnson H, Eyers CE, Eyers PA, Beynon RJ, Gaskell SJ.Rigorous determination of the stoichiometry of protein phosphorylation using mass spectrometry. J Am Soc Mass Spectrom. 2009 Dec;20(12):2211-20. [PubMed]
  6. Brownridge P, Lawless C, Payapilly AB, Lanthaler K, Holman SW, Harman VM, Grant CM, Beynon RJ, Hubbard SJ. Quantitative analysis of chaperone network throughput in budding yeast. Proteomics. 2013 Apr;13(8):1276-91. [PubMed]
  7. Carroll KM, Simpson DM, Eyers CE, Knight CG, Brownridge P, Dunn WB, Winder CL, Lanthaler K, Pir P, Malys N, Kell DB, Oliver SG, Gaskell SJ, Beynon RJ. Absolute quantification of the glycolytic pathway in yeast: deployment of a complete QconCAT approach. Mol Cell Proteomics. 2011 Dec;10(12):M111.007633. [PubMed]
  8. Southworth PM, Hyde JE, Sims PF. A mass spectrometric strategy for absolute quantification of Plasmodium falciparum proteins of low abundance. Malar J. 2011 Oct 25;10:315. [PubMed]
  9. Castro-Borges W, Simpson DM, Dowle A, Curwen RS, Thomas-Oates J, Beynon RJ, Wilson RA. Abundance of tegument surface proteins in the human blood fluke Schistosoma mansoni determined by QconCAT proteomics. J Proteomics. 2011 Aug 24;74(9):1519-33. [PubMed]
  10. Brownridge P, Holman SW, Gaskell SJ, Grant CM, Harman VM, Hubbard SJ, Lanthaler K, Lawless C, O’Cualain R, Sims P, Watkins R, Beynon RJ. Global absolute quantification of a proteome: Challenges in the deployment of a QconCAT strategy. Proteomics. 2011 Aug;11(15):2957-70. [PubMed]
  11. Lawless C, Holman SW, Brownridge P, Lanthaler K, Harman VM, Watkins R, Hammond DE, Miller RL, Sims PF, Grant CM, Eyers CE, Beynon RJ, Hubbard SJ. Direct and Absolute Quantification of over 1800 Yeast Proteins via Selected Reaction Monitoring. Mol Cell Proteomics. 2016 Apr;15(4):1309-22. [Pubmed]
  12. Peffers MJ, Beynon RJ, Clegg PD. Absolute quantification of selected proteins in the human osteoarthritic secretome. Int J Mol Sci. 2013 Oct 15;14(10):20658-81. [PubMed]
  13. Mackenzie RJ, Lawless C, Holman SW, Lanthaler K, Beynon RJ, Grant CM, Hubbard SJ, Eyers CE. Absolute protein quantification of the yeast chaperome under conditions of heat shock. Proteomics. 2016 Jun 2; [PubMed]

 

Contact us to find out more about the implementation of custom-made QconCATs in your research project or benefit from our experience by confiding us with a full service quantification project.

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