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Mechanism of Coomassie brilliant blue G-250 binding to proteins: a hydrophobic assay for nanogram quantities of proteins

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Abstract

We investigated the mechanism of Coomassie brilliant blue G-250 (CBB) binding to proteins in order to develop a protein assay with the maximum possible sensitivity. We found that the neutral ionic species of CBB binds to proteins by a combination of hydrophobic interactions and heteropolar bonding with basic amino acids. On the basis of these findings, we developed a very sensitive hydrophobic assay for proteins (at the nanogram level) using the hydrophobic reagents ammonium sulfate and trichloroacetic acid under pH conditions that increase neutral species concentration in the assay reagent in order to enhance the binding of more CBB dye molecules per protein molecule than in previous CBB-based assays.

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Acknowledgements

This work was supported financially by the Greek Ministry of Education, University of Patras, Greece. K.G. thanks the Cultural Institute of Moral and Social Education, Athens, Greece, for supporting him financially.

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Authors and Affiliations

  1. Department of Biology, Section of Genetics, Cell Biology and Development, University of Patras, 26100, Patras, Greece

    Christos D. Georgiou, Konstantinos Grintzalis & Ioannis Papapostolou

  2. Department of Greenhouse Crops and Floriculture, Technological Institute of Mesologgi, 30200, Nea Ktiria, Mesologgi, Greece

    George Zervoudakis

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  1. Christos D. Georgiou
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  2. Konstantinos Grintzalis
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  3. George Zervoudakis
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  4. Ioannis Papapostolou
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Correspondence to Christos D. Georgiou.

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Georgiou, C.D., Grintzalis, K., Zervoudakis, G. et al. Mechanism of Coomassie brilliant blue G-250 binding to proteins: a hydrophobic assay for nanogram quantities of proteins. Anal Bioanal Chem 391, 391–403 (2008). https://doi.org/10.1007/s00216-008-1996-x

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  • DOI: https://doi.org/10.1007/s00216-008-1996-x

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