QconCAT

What is a QconCAT?

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

 

QconCAT 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
  • Rutten MGS, Derks TGJ, Huijkman NCA, Bos T, Kloosterhuis NJ, van de Kolk KCWA, Wolters JC, Koster MH, Bongiovanni J, Thomas RE, de Bruin A, van de Sluis B, and Oosterveer MH Modeling Phenotypic Heterogeneity of Glycogen Storage Disease Type 1a Liver Disease in Mice by Somatic CRISPR/CRISPR-associated protein 9– Mediated Gene Editing
    HEPATOLOGY, VOL. 74, NO. 5, 2021[Pubmed]
  • Vasilogianni AM, Al-Majdoub ZM, Achour B, Peters SA, Rostami-Hodjegan A, Barber J; Proteomics of Colorectal Cancer Liver Metastasis: a Quantitative Focus on Drug Elimination and Pharmacodynamics Effects
    Br J Clin Pharmacol. 2021 Oct 1.[Pubmed]
  • Heida A, Gruben N, Catrysse L, Koehorst M, Koster M, Kloosterhuis NJ, Gerding A, Havinga R, Bloks VM, Bongiovanni L, Wolters JC, van Dijk T, van Loo G, de Bruin A, Kuipers F, Koonen DPY, van de Sluis B The hepatocyte IKK:NF-kB axis promotes liver steatosis by stimulating de novo lipogenesis and cholesterol synthesis
    MOLECULAR METABOLISM 54 (2021)[Pubmed]
  • El-Khateeb E, Achour B, Al-Majdoub ZM, Barber J, Rostami-Hodjegan A; Non-uniformity of Changes in Drug-Metabolizing Enzymes and Transporters in Liver Cirrhosis: Implications for Drug Dosage Adjustment.
    Mol Pharm. 2021 Sep 6;18(9):3563-3577.[Pubmed]
  • Vieira-Lara MA, Dommerholt MB, Zhang W, Blankestijn M, Wolters JC, Abegaz F, Gerding A, van der Veen YT, Thomas R, van Os RP, Reijngoud DJ, Jonker JW, Kruit JK and Bakker BM Age-related susceptibility to insulin resistance arises from a combination of CPT1B decline and lipid overload
    BMC Biology (2021) 19:154.[Pubmed]
  • Alejandro Sánchez Brotons, Jonatan O. Eriksson, Marcel Kwiatkowski, Justina C. Wolters, Ido P. Kema, Andrei Barcaru, Folkert Kuipers, Stephan J. L. Bakker, Rainer Bischoff, Frank Suits, and Péter Horvatovich Pipelines and Systems for Threshold-Avoiding Quantification of LC−MS/MS Data
    Anal. Chem. 2021, 93, 11215−11224.[Pubmed]
  • Joanne A. Hoogerland, Fabian Peeks, Brenda S. Hijmans, Justina C. Wolters, Sander Kooijman, Trijnie Bos, Aycha Bleeker, Theo H. van Dijk, Henk Wolters, Albert Gerding, Karen van Eunen, Rick Havinga, Amanda C. M. Pronk, Patrick C. N. Rensen, Gilles Mithieux, Fabienne Rajas, Folkert Kuipers, Dirk-Jan Reijngoud, Terry G. J. Derks, Maaike H. Oosterveer Impaired Very-Low-Density Lipoprotein catabolism links hypoglycemia to hypertriglyceridemia in Glycogen Storage Disease type Ia
    J Inherit Metab Dis. 2021;44:879–892.[Pubmed]
  • Jan Freark de Boer, Hilde D. de Vries, Anna Palmiotti, Rumei Li, Marwah Doestzada, Joanne A. Hoogerland, Jingyuan Fu, Anouk M. La Rose, Marit Westerterp, Niels L. Mulder, Milaine V. Hovingh, Martijn Koehorst, Niels J. Kloosterhuis, Justina C. Wolters, Vincent W. Bloks, Joel T. Haas, David Dombrowicz, Bart Staels, Bart van de Sluis, and Folkert Kuipers Cholangiopathy and Biliary Fibrosis in Cyp2c70-Deficient Mice Are Fully Reversed by Ursodeoxycholic Acid
    Cell Mol Gastroenterol Hepatol 2021;11:1045–1069.[Pubmed]
  • Eman El-Khateeb, Zubida M. Al-Majdoub, Amin Rostami-Hodjegan, Jill Barber, and Brahim Achour Proteomic Quantification of Changes in Abundance of Drug-Metabolizing Enzymes and Drug Transporters in Human Liver Cirrhosis: Different Methods, Similar Outcomes
    Drug Metabolism and Disposition May 27, 2021.[Pubmed]
  • Bart Van Puyvelde, Katleen Van Uytfanghe, Olivier Tytgat, Laurence Van Oudenhove, Ralf Gabriels, Robbin Bouwmeester, Simon Daled, Tim Van Den Bossche, Pathmanaban Ramasamy, Sigrid Verhelst, Laura De Clerck, Laura Corveleyn, Sander Willems, Nathan Debunne, Evelien Wynendaele, Bart De Spiegeleer, Peter Judak, Kris Roels, Laurie De Wilde, Peter Van Eenoo, Tim Reyns, Marc Cherlet, Emmie Dumont, Griet Debyser, Ruben t’Kindt, Koen Sandra, Surya Gupta, Nicolas Drouin, Amy Harms, Thomas Hankemeier, Donald J. L. Jones, Pankaj Gupta, Dan Lane, Catherine S. Lane, Said El Ouadi, Jean-Baptiste Vincendet, Nick Morrice, Stuart Oehrle, Nikunj Tanna, Steve Silvester, Sally Hannam, Florian C. Sigloch, Andrea Bhangu-Uhlmann, Jan Claereboudt, N. Leigh Anderson, Morteza Razavi, Sven Degroeve, Lize Cuypers, Christophe Stove, Katrien Lagrou, Geert A. Martens, Dieter Deforce, Lennart Martens, Johannes P. C. Vissers, and Maarten Dhaenens; Cov-MS: A Community-Based Template Assay for Mass-Spectrometry-Based Protein Detection in SARS-CoV-2 Patients
    JACS Au 2021, 1, 6, 750–765.[Pubmed]
  • Reichermeier KM, Straube R, Reitsma JM, Sweredoski MJ, Rose CM, Moradian A, den Besten W, Hinkle T, Verschueren E, Petzold G, Thomä NH, Wertz IE, Deshaies RJ, Kirkpatrick DS; PIKES Analysis Reveals Response to Degraders and Key Regulatory Mechanisms of the CRL4 Network.
    Mol Cell. 2020 Jan 15.[Pubmed]
  • 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]

 

Examples for QconCAT application

Original publications introducing the QconCAT method
  • 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]
  • 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]
Large scale quantification
  • 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]
  • 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]
  • 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]
  • 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]
  • 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]
Stoichiometry
  • 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]
  • 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]
Diagnostics
  • Bart Van Puyvelde, Katleen Van Uytfanghe, Olivier Tytgat, Laurence Van Oudenhove, Ralf Gabriels, Robbin Bouwmeester, Simon Daled, Tim Van Den Bossche, Pathmanaban Ramasamy, Sigrid Verhelst, Laura De Clerck, Laura Corveleyn, Sander Willems, Nathan Debunne, Evelien Wynendaele, Bart De Spiegeleer, Peter Judak, Kris Roels, Laurie De Wilde, Peter Van Eenoo, Tim Reyns, Marc Cherlet, Emmie Dumont, Griet Debyser, Ruben t’Kindt, Koen Sandra, Surya Gupta, Nicolas Drouin, Amy Harms, Thomas Hankemeier, Donald J. L. Jones, Pankaj Gupta, Dan Lane, Catherine S. Lane, Said El Ouadi, Jean-Baptiste Vincendet, Nick Morrice, Stuart Oehrle, Nikunj Tanna, Steve Silvester, Sally Hannam, Florian C. Sigloch, Andrea Bhangu-Uhlmann, Jan Claereboudt, N. Leigh Anderson, Morteza Razavi, Sven Degroeve, Lize Cuypers, Christophe Stove, Katrien Lagrou, Geert A. Martens, Dieter Deforce, Lennart Martens, Johannes P. C. Vissers, and Maarten Dhaenens; Cov-MS: A Community-Based Template Assay for Mass-Spectrometry-Based Protein Detection in SARS-CoV-2 Patients
    JACS Au 2021, 1, 6, 750–765.[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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