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On the utility of fluorescence-detection analytical ultracentrifugation in probing biomolecular interactions in complex solutions: a case study in milk

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Abstract

β-Lactoglobulin is the most abundant protein in the whey fraction of ruminant milks, yet is absent in human milk. It has been studied intensively due to its impact on the processing and allergenic properties of ruminant milk products. However, the physiological function of β-lactoglobulin remains unclear. Using the fluorescence-detection system within the analytical ultracentrifuge, we observed an interaction involving fluorescently labelled β-lactoglobulin in its native environment, i.e. cow and goat milk, for the first time. Co-elution experiments support that these β-lactoglobulin interactions occur naturally in milk and provide evidence that the interacting partners are immunoglobulins, while further sedimentation velocity experiments confirm that an interaction occurs between these molecules. The identification of these interactions, made possible through the use of fluorescence-detected analytical ultracentrifugation, provides possible clues to the long debated physiological function of this abundant milk protein.

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Abbreviations

FITC:

Fluorescein isothiocyanate

IgG:

Immunoglobulin G

IgA:

Immunoglobulin A

IgM:

Immunoglobulin M

AUC:

Analytical ultracentrifugation

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Acknowledgements

We acknowledge the following for funding support, in part: (1) the New Zealand Ministry of Business, Innovation and Employment, (2) the New Zealand Royal Society Marsden Fund, (3) the Biomolecular Interaction Centre, University of Canterbury, and (4) The Riddet Institute.

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Authors and Affiliations

  1. Biomolecular Interaction Centre, School of Biological Sciences, University of Canterbury, Christchurch, New Zealand

    Jennifer M. Crowther, Geoffrey B. Jameson & Renwick C. J. Dobson

  2. The Riddet Institute, Massey University, Palmerston North, New Zealand

    Jennifer M. Crowther, Geoffrey B. Jameson & Renwick C. J. Dobson

  3. Food and Bio-Based Products, AgResearch Limited, Ruakura Research Centre, Hamilton, New Zealand

    Marita Broadhurst & Alison J. Hodgkinson

  4. Center To Advance Molecular Interaction Science, University of New Hampshire, Durham, NH, USA

    Thomas M. Laue

  5. School of Fundamental Sciences, Massey University, Palmerston North, New Zealand

    Geoffrey B. Jameson

  6. Department of Biochemistry and Molecular Biology, Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, Melbourne, VIC, Australia

    Renwick C. J. Dobson

  7. On-Farm R&D, Farm Source, Fonterra Co-Operative Group, Hamilton, 3200, New Zealand

    Alison J. Hodgkinson

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  1. Jennifer M. Crowther
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  2. Marita Broadhurst
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Correspondence to Jennifer M. Crowther, Alison J. Hodgkinson or Renwick C. J. Dobson.

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Special Issue: Analytical Ultracentrifugation 2019.

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Crowther, J.M., Broadhurst, M., Laue, T.M. et al. On the utility of fluorescence-detection analytical ultracentrifugation in probing biomolecular interactions in complex solutions: a case study in milk. Eur Biophys J 49, 677–685 (2020). https://doi.org/10.1007/s00249-020-01468-3

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