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Temperature denaturation and aggregation of a multi-domain protein (IgG1) investigated with an array of complementary biophysical methods

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Abstract

Proteins are used as drugs against different pathologies because of their potential specificity of action with fewer side effects. However, their production and successful storage imposes a greater challenge compared to small molecule drugs. Though the determination of protein thermal stability is commonly used to find the optimum storage conditions for biopharmaceuticals, a multi-technique approach should be applied more often when investigating complex systems to understand the structure of the species that contribute to the different transitions, thereby gaining insight about the processes of both unfolding and aggregation. This knowledge is crucial for identifying those conformational changes which are likely to lead to aggregation/degradation allowing a more rational approach to biopharmaceutical production and formulation. This is particularly important in the case of multi-domain proteins, such as IgGs, which can undergo multiple transitions due to independent unfolding of the domains. In this work, we have followed the thermal denaturation of a monoclonal antibody by using different biophysical techniques with complementary strengths, providing an example of how the information gathered suggests a way to intervene to stabilise the wanted conformation (monomeric protein). Indeed, in this particular case, an optimisation of storage conditions based on only thermal stability studies would have led to the stabilisation of an undesired product, a population of low molecular weight oligomers.

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Acknowledgments

We thank the UK Department of Business Innovation and Skills for funding the work and GSK for the immunoglobulin sample. We thank Alex Knight, Anna Hills, Adrian Horgan, Paulina Rakowska and Max Ryadnov for reviewing the manuscript and Michael Molloy for sourcing the sample.

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Author notes

  1. T. Garfagnini

    Present address: Institute of Chemistry, The Hebrew University of Jerusalem, Safra Campus, Givat Ram, 91904, Jerusalem, Israel

Authors and Affiliations

  1. Biotechnology Group, National Physical Laboratory, Hampton Road, Teddington, Middlesex, TW11 0LW, UK

    E. Cerasoli, J. Ravi, T. Garfagnini & G. E. Tranter

  2. Materials - Composites / Thermal Analysis, National Physical Laboratory, Hampton Road, Teddington, Middlesex, TW11 0LW, UK

    S. Gnaniah

  3. Chiralabs Limited, Begbroke Centre for Innovation and Enterprise, Begbroke Science Park, Begbroke Hill, Woodstock Road, Begbroke, Oxfordshire, OX5 1PF, UK

    D. le Pevelen & G. E. Tranter

Authors
  1. E. Cerasoli
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  2. J. Ravi
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  3. T. Garfagnini
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  4. S. Gnaniah
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  5. D. le Pevelen
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  6. G. E. Tranter
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Corresponding author

Correspondence to E. Cerasoli.

Additional information

Published in the topical collection Analysis of Biological Therapeutic Agents and Biosimilars with guest editor Karen Phinney.

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Cerasoli, E., Ravi, J., Garfagnini, T. et al. Temperature denaturation and aggregation of a multi-domain protein (IgG1) investigated with an array of complementary biophysical methods. Anal Bioanal Chem 406, 6577–6586 (2014). https://doi.org/10.1007/s00216-014-7970-x

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  • DOI: https://doi.org/10.1007/s00216-014-7970-x

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