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    Small and Stable Peptidic PEGylated Quantum Dots to Target Polyhistidine-Tagged Proteins with Controlled Stoichiometry

    Year: 2009

    Journal: J. Am. Chem. Soc., 2009, 131 (41), pp 14738–14746, 20100827

    Authors: Dif A. 1, Boulmedais F. 1 3, Pinot M. 2, Roullier V. 1, Baudy-Floc’h M. 1, Coquelle F.M. 4, Clarke S. 6, Neveu P. 5 *,  Vignaux F. 7, Le Borgne R. 7, Dahan M. 6, Gueroui Z. 2, Marchi-Artzner V. *1

    Last authors: Valerie Marchi-Artzner

    Organizations: 1 Université de Rennes 1, CNRS UMR 6226, Sciences Chimiques de Rennes. 2 Université de Rennes 1, CNRS UMR 6251, Institut de Physique de Rennes. 3 Institut Charles Sadron, CNRS UPR 22. 4 Université de Rennes 1, CNRS UMR 6062, IFR 140. 5 Ecole Normale Supérieure, CNRS UMR 8640, Département de Chimie. 6 Laboratoire Kastler Brossel, Ecole Normale Supérieure, CNRS UMR 8552, Université Pierre et Marie Curie. 7 Université de Rennes 1, CNRS UMR 6061, Faculté de Médecine. * Present address: Kavli Institute for Theoretical Physics, University of California at Santa Barbara.

    Country: France

    The use of the semiconductor quantum dots (QD) as biolabels for both ensemble and single-molecule tracking requires the development of simple and versatile methods to target individual proteins in a controlled manner, ideally in living cells. To address this challenge, we have prepared small and stable QDs (QD-ND) using a surface coating based on a peptide sequence containing a tricysteine, poly(ethylene glycol) (PEG), and an aspartic acid ligand. These QDs, with a hydrodynamic diameter of 9 ± 1.5 nm, can selectively bind to polyhistidine-tagged (histag) proteins in vitro or in living cells. We show that the small and monodisperse size of QD-ND allows for the formation of QD-ND/histag protein complexes of well-defined stoichiometry and that the 1:1 QD/protein complex can be isolated and purified by gel electrophoresis without any destabilization in the nanomolar concentration range. We also demonstrate that QD-ND can be used to specifically label a membrane receptor with an extracellular histag expressed in living HeLa cells. Here, cytotoxicity tests reveal that cell viability remains high under the conditions required for cellular labeling with QD-ND. Finally, we apply QD-ND complexed with histag end binding protein-1 (EB1), a microtubule associated protein, to single-molecule tracking in Xenopus extracts. Specific colocalization of QD-ND/EB1 with microtubules during the mitotic spindle formation demonstrates that QD-ND and our labeling strategy provide an efficient approach to monitor the dynamic behavior of proteins involved in complex biological functions.