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Application of Quartz Crystal Microbalance with Dissipation Monitoring Technology for Studying Interactions of Poxviral Proteins with Their Ligands

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Vaccinia Virus and Poxvirology

Part of the book series: Methods in Molecular Biology ((MIMB,volume 890))

Abstract

Poxviruses are one of the most complex of animal viruses and encode for over 150 proteins. The interactions of many of the poxviral-encoded proteins with host proteins, as well as with other proteins, such as transcription complexes, have been well characterized at the qualitative level. Some have also been characterized quantitatively by two hybrid systems and surface plasmon resonance approaches. Presented here is an alternative approach that can enable the understanding of complex interactions with multiple ligands. The example given is that of vaccinia virus complement control protein (VCP). The complement system forms the first line of defense against microorganisms and a failure to appropriately regulate it is implicated in many inflammatory disorders, such as traumatic brain injury, Alzheimer’s disease (AD), and rheumatoid arthritis. The complement component C3 is central to the complement activation. Complement regulatory proteins, capable of binding to the central complement component C3, may therefore effectively be employed for the treatment and prevention of these disorders. There are many biochemical and/or immunoassays available to study the interaction of proteins with complement components. However, protocols for many of them are time consuming, and not all assays are useful for multiple screening. In addition, most of these assays may not give information regarding the nature of binding, the number of molecules interacting with the complement component C3, as well as kinetics of binding. Some of the assays may require labeling which may induce changes in protein confirmation. We report a protocol for an assay based on quartz crystal microbalance with dissipation monitoring (QCM-D) technology, which can effectively be employed to study poxviral proteins for their ability to interact with their ligand. A protocol was developed in our laboratories to study the interaction of VCP with the complement component C3 using Q-sense (D-300), equipment based on QCM-D technology. The protocol can also be used as a prototype for studying both proteins and small-sized compounds (for use as anti-poxvirals) for their ability to interact with and/or inhibit the activity of their ligands.

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Author information

Authors and Affiliations

  1. Division of Anatomical Pathology, Department of Clinical Laboratory sciences, Faculty of Health Sciences, University of Cape Town, Cape Town, South Africa

    Amod P. Kulkarni & Lauriston A. Kellaway

  2. Kotwal Bioconsulting, LLC, Louisville, KY, USA

    Girish J. Kotwal

  3. InFlaMed Inc, Louisville, KY, USA

    Girish J. Kotwal

  4. Department of Microbiology and Biochemistry, University of Medicine and Health Sciences, Saint Kitts, West Indies

    Girish J. Kotwal

Authors
  1. Amod P. Kulkarni
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  2. Lauriston A. Kellaway
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  3. Girish J. Kotwal
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Corresponding authors

Correspondence to Lauriston A. Kellaway or Girish J. Kotwal .

Editor information

Editors and Affiliations

  1. School of Medicine and the Philadelphia, Division of Infectious Disease, University of Pennsylvania, Johnson Pavilion 502, Philadelphia, 19104-6073, Pennsylvania, USA

    Stuart N. Isaacs

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Kulkarni, A.P., Kellaway, L.A., Kotwal, G.J. (2012). Application of Quartz Crystal Microbalance with Dissipation Monitoring Technology for Studying Interactions of Poxviral Proteins with Their Ligands. In: Isaacs, S. (eds) Vaccinia Virus and Poxvirology. Methods in Molecular Biology, vol 890. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-876-4_17

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  • DOI: https://doi.org/10.1007/978-1-61779-876-4_17

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-61779-875-7

  • Online ISBN: 978-1-61779-876-4

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