Metal-ligand complexes as a new class of long-lived fluorophores for protein hydrodynamics

Abstract

We describe the use of asymmetric Ru-ligand complexes as a new class of luminescent probes that can be used to measure rotational motions of proteins. These complexes are known to display luminescent lifetimes ranging from 10 to 4000 ns. In this report, we show that the asymmetric complex Ru(bpy)2(dcbpy) (PF6)2 displays a high anisotropy value when excited in the long wavelength absorption band. For covalent linkage to proteins, we synthesized the N-hydroxy succinimide ester of this metal-ligand complex. To illustrate the usefulness of these probes, we describe the intensity and anisotropy decays of [Ru(bpy)2(dcbpy)] when covalently linked to human serum albumin, concanavalin A (ConA), human immunoglobulin G (IgG), and Ferritin, and measured in solutions of increased viscosity. These data demonstrate that the probes can be used to measure rotational motions on the 10 ns to 1.5 microseconds timescale, which so far has been inaccessible using luminescence methods. The present probe [Ru(bpy)2(dcbpy)] can be regarded as the first of a class of metal-ligand complexes, each with different chemical reactivity and spectral properties, for studies of macromolecular dynamics.

Advances in Fluorescence Spectroscopy: Multi-Photon Excitation, Engineered Proteins, Modulation Sensing and Microsecond Rhenium Metal-Ligand Complexes

“…This property provides information about rotational motions from the steady-state or time-resolved anisotropies. We found that the ruthenium complexes display polarized emission, allowing their use as anisotropy probes [63][64][65][66][67]. For these complexes the emission is from a metal-ligand charge transfer (MLCT) state.…”

Wide-field TCSPC: methods and applications

“…The long lifetime increases the sensitivity by giving the probe more time to interact with the environment, and it allows the short-lived autofluorescence from the sample to be discarded. Phosphorescence anisotropy measurements can also be used to study the rotational diffusion of large proteins, whose movement is too slow to be measured with fast fluorescence decays [182,183]. Besides life sciences, PLIM has been used to study air-flow and pressure in aerodynamic studies [184], and for temperature measurements in industrial thermometry applications [185].…”

Electrochemical and Electrochemiluminescence Determination of Cancer Cells Based on Aptamers and Magnetic Beads

“…[38,39] The as-prepared [Ru-A C H T U N G T R E N N U N G (bpy) 2 Cl 2 ] (0.16 g), NaHCO 3 A C H T U N G T R E N N U N G (0.16 g), and 2,2'-bipyridine-4,4'-dicarboxylic acid (0.12 g) were added to a solution of methanol and H 2 O solution (20 mL; 4:1 v/v) and the stirred solution was heated for 4 h. The solution was cooled in an ice bath for 7 h, and the pH of the solution was adjusted with hydrochloric acid to pH 4.4. The formed precipitate was filtered and washed with methanol.…”