Post-protein binding metal-mediated coupling of an acridine orange-based fluorophore.
The HEW lysozyme (Lys) and the fac-[Re(CO)(3)(H(2)O)(3)](+) complex (1) are used as a simple model system for the description of a new approach to the labelling polypeptides with fluorescent tags. The strategy takes advantage of the reaction of an acridine orange-based fluorophore (AO) with the non-native metal fragment 1 hybridized on the enzyme. A synthetic methodology for the quantitative metallation of the protein is first described and it is then shown that the exogenous metal complex can be exploited for the coupling of the fluorescent probe. All Lys-derived species were characterized by various spectroscopic techniques. It is shown that the approach does not significantly alter the activity of the final fluorescent metallo-protein conjugate (Lys2). The accumulation of Lys2 on Micrococcus lysodeikticus bacteria was observed via confocal laser scanning microscopy.
Institute of Inorganic Chemistry, University of Zürich, Winterthurerstrasse 190, CH-8057 Zürich, Switzerland.
Giuseppe
G
Santoro
Olivier
O
Blacque
Fabio
F
Zobi
D000165:Acridine Orange / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid; D005456:Fluorescent Dyes / Q000737:chemistry*; D008837:Micrococcus / Q000378:metabolism; D018613:Microscopy, Confocal; D008956:Models, Chemical; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary
D005456:Fluorescent Dyes; D010455:Peptides; D000165:Acridine Orange