Fluorescence Microscopy of Single Liposomes with Incorporated Pigment-Proteins

Marijonas Tutkus, Parveen Akhtar, Jevgenij Chmeliov, Fanni Görföl, Gediminas Trinkunas, P. Lambrev, Leonas Valkunas

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4 Citations (Scopus)

Abstract

Reconstitution of transmembrane proteins into liposomes is a widely used method to study their behavior under conditions closely resembling the natural ones. However, this approach does not allow precise control of the liposome size, reconstitution efficiency, and the actual protein-to-lipid ratio in the formed proteoliposomes, which might be critical for some applications and/or interpretation of data acquired during the spectroscopic measurements. Here, we present a novel strategy employing methods of proteoliposome preparation, fluorescent labeling, purification, and surface immobilization that allow us to quantify these properties using fluorescence microscopy at the single-liposome level and for the first time apply it to study photosynthetic pigment-protein complexes LHCII. We show that LHCII proteoliposome samples, even after purification with a density gradient, always contain a fraction of nonreconstituted protein and are extremely heterogeneous in both protein density and liposome sizes. This strategy enables quantitative analysis of the reconstitution efficiency of different protocols and precise fluorescence spectroscopic study of various transmembrane proteins in a controlled nativelike environment.

Original languageEnglish
Pages (from-to)14410-14418
Number of pages9
JournalLangmuir
Volume34
Issue number47
DOIs
Publication statusPublished - Nov 27 2018

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ASJC Scopus subject areas

  • Materials Science(all)
  • Condensed Matter Physics
  • Surfaces and Interfaces
  • Spectroscopy
  • Electrochemistry

Cite this

Tutkus, M., Akhtar, P., Chmeliov, J., Görföl, F., Trinkunas, G., Lambrev, P., & Valkunas, L. (2018). Fluorescence Microscopy of Single Liposomes with Incorporated Pigment-Proteins. Langmuir, 34(47), 14410-14418. https://doi.org/10.1021/acs.langmuir.8b02307