Luminescence quenching by long range electron transfer: A probe of protein clustering and conformation at the cell surface

Janos Matko, Attila Jenei, Taiyin Wei, Michael Edidin

Research output: Contribution to journalArticlepeer-review

Abstract

Quenching of luminescence from fluorescent and phosphorescent probes by nitroxide spin labels with a long range electron transfer (LRET) mechanism (44,45) has been tested as a tool to monitor association/clustering and conformational changes of cell surface proteins. The membrane proteins were labeled with monoclonal antibodies or Fab fragments conjugated with luminescent probes or water‐soluble nitroxide spin labels. The method was tested as a probe of 3 different aspects of protein‐protein association involving class I MHC molecules: (1) interaction between the heavy and light chains of the MHC molecules, (2) clustering, self‐association of MHC molecules, (3) proximity of MHC molecules to transferrin receptors of fibroblasts or surface immunoglobulin molecules of B lymphoblasts. The extent of quenching upon increasing the fractional density of the quencher was sensitive for protein association in accordance with earlier immunoprecipitation and flow cytometric Förster‐type energy transfer (FCET) data obtained on the same cells. These data suggest that the LRET quenching can be used as intra‐ or intermolecular ruler in a 0.5–2.5 nm distance range. This approach is simpler (measurements only on donor side) and faster than many other experimental techniques in screening physical association or conformational changes of membrane proteins by means of spectrofluorimetry, flow cytometry, or microscope based imaging. © 1995 Wiley‐Liss, Inc.

Original languageEnglish (US)
Pages (from-to)191-200
Number of pages10
JournalCytometry
Volume19
Issue number3
DOIs
StatePublished - Mar 1 1995

Keywords

  • Fluorescence quenching
  • clusters
  • electron transfer
  • flow cytometry
  • protein association

ASJC Scopus subject areas

  • Pathology and Forensic Medicine
  • Biophysics
  • Hematology
  • Endocrinology
  • Cell Biology

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