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Tracing Lysozyme-Lipid Interactions with Long-Wavelength Squaraine Dyes

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Abstract

The applicability of the two newly commercial available squaraine labels Square-670-NHS and Seta-635-NHS to exploring protein-lipid interactions has been evaluated. The labels were conjugated to lysozyme (Lz) (squaraine-lysozyme conjugates below referred to as Square-670-Lz and Seta-635-Lz), a structurally well-characterized small globular protein displaying the ability to interact both, electrostatically and hydrophobically with lipids. The lipid component of the model systems was represented by lipid vesicles composed of zwitterionic lipids egg yolk phosphatidylcholine (PC) and 1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine (SOPC), and their mixtures with anionic lipids either beef heart cardiolipin (CL) or 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG), respectively. Fluorescence intensity of Square-670-Lz was found to decrease upon association with lipid bilayer, while the fluorescence intensity of Seta-635-Lz displayed more complex behavior depending on lipid-to-protein molar ratio. Covalent coupling of squaraine labels to lysozyme exerts different influence on the properties of dye-protein conjugate. It was suggested that Square-670-NHS covalent attachment to Lz molecule enhances protein propensity for self-association, while squaraine label Seta-635-NHS is sensitive to different modes of lysozyme-lipid interactions—within the L:P range 6–11, when hydrophobic protein-lipid interactions are predominant, an aggregation of membrane-bound protein molecules takes place, thereby decreasing the fluorescence intensity of Seta-635-Lz. At higher L:P values (from 22 to 148) when electrostatic interactions are enhanced fluorescence intensity of Seta-635-Lz increases with increasing lipid concentrations.

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Acknowledgments

This work was supported by grant from the Sigrid Juselius Foundation.

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Authors and Affiliations

  1. Department of Biological and Medical Physics, V.N. Karazin Kharkov National University, 4 Svobody Sq., 32-90 Geroyev Truda St., Kharkov, 61146, Ukraine

    Valeriya M. Ioffe & Galyna P. Gorbenko

  2. Helsinki Biophysics and Biomembrane Group, Institute of Biomedicine, University of Helsinki, Haartmaninkatu 8, P.O. Box 63, FIN-00014, Helsinki, Finland

    P. K. J. Kinnunen

  3. STC “Institute for Single Crystals” of the National Academy of Sciences of Ukraine, 60 Lenin Ave., Kharkov, 61001, Ukraine

    Anatoliy L. Tatarets, Olga S. Kolosova & Leonid D. Patsenker

  4. SETA BioMedicals, LLC, 2014 Silver Ct East, Urbana, IL, 61801, USA

    Ewald A. Terpetschnig

Authors
  1. Valeriya M. Ioffe
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  2. Galyna P. Gorbenko
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  3. P. K. J. Kinnunen
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  4. Anatoliy L. Tatarets
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  5. Olga S. Kolosova
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  6. Leonid D. Patsenker
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  7. Ewald A. Terpetschnig
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Correspondence to Valeriya M. Ioffe.

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Ioffe, V.M., Gorbenko, G.P., Kinnunen, P.K.J. et al. Tracing Lysozyme-Lipid Interactions with Long-Wavelength Squaraine Dyes. J Fluoresc 17, 65–72 (2007). https://doi.org/10.1007/s10895-006-0142-4

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  • DOI: https://doi.org/10.1007/s10895-006-0142-4

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