Selective fluorescent chemosensors and iodinated contrast agents : biological evaluation for cellular zinc detection, microdamage detection and imaging

THESIS 7554This thesis entitled "Selective Fluorescent Chemosensors and lodinated Contrast Agents: Biological Evaluation for Cellular Zinc detection, Microdamage Detection and Imaging" has been divided into six Chapters. Chapter 1 highlights advances made in the field of fluorescent chemosensors. Examples discussed have been critically evaluated in terms of their photophysical properties, selectivity, sensitivity and binding ability. To introduce the relevance of supramolecular systems for microdamage detection in bone, a brief introduction to the characteristics of bone and X-rays are also provided. Chapter 2 presents in detail the design, synthesis and photophysical evaluation of two water soluble and selective Zn(II) fluoroprobes 92 and 93. The key feature of these sensors is their stability under simulated physiological conditions. The fluorescence titrations of 92 demonstrated its ability to detect nanomolar free Zn(II). Preliminarily studies carried out using pancreatic tissue have shown the ability of 92 to label the areas, which contain chelatable zinc. Chapter 3 presents in details two aqueous soluble, anthracene based fluorescent PET sensors 125 and 126 for selective recognition of Zn(II) and Cd(II). The evaluation of the photophysical response o f these sensors demonstrated their ability to distinguish between Zn(II) and Cd(II). Chapter 4 details a critical investigation of microdamage in bone using histological methods, SEM and EDXA. A set of fluoroprobes, 135, 136, 138, 140, 141, 142, 143, 92, 93 and 125 were used to investigate the selective labelling of the microcrack in the bone. The scratched and unscratched surfaces of the bone were studied using Raman spectroscopy, SEM and EDXA. The selective labelling of scratches observed can be attributed to the interaction of dyes via binding at the free lattice sites, ionic interactions with the free lattice sites or incorporation in the broken lattice sites

Luminescent sensing

The development of luminescent sensing has been central to the field of supramolecular chemistry. Herein, we give a some insight into some of the developments that have taken place in recent times, focusing most of our effort on the discussion of luminescent sensors for cations, anions, and neutral molecules, by presenting a selected number of examples from this fast, broad, and ever expanding research field. We also demonstrate how the use of luminescent sensors has helped in the development of contrast and imaging agents, and in particular show examples of their use in cellular imaging and sensing within competitive biological environments

DprE1-DprE2 interaction analysis.

<p>(a) The stereo view of the binding surface of the complex where DprE1 and DprE2 are represented by red and green respectively. (b) Cartoon representation of the complex showing interface residues involved in H-bonding where grey and green represents carbon atoms of DprE1 and DprE2 respectively. Oxygen ayoms are coloured red whereas nitrogen atoms are blue. The dashed line represents the H-bond network between residues of DprE1 and DprE2.</p

PrincipalComponent analysis of DprE1 and DprE2.

<p>Covariance matrix of the fluctuations of the Cα atoms of (a) DprE1 and (b) DprE2 during the simulationswith positive (red) and negative (blue) motions. (c) and (d) represent subspace overlap for the first 10 eigenvectors of the first half (0–25 ns) with all the eigenvectors of the second half (25–50 ns) of the simulation for DprE1 and DprE2 respectively.</p

Structure, Dynamics, and Interaction of <i>Mycobacterium tuberculosis</i> (<i>Mtb</i>) DprE1 and DprE2 Examined by Molecular Modeling, Simulation, and Electrostatic Studies

<div><p>The enzymes decaprenylphosphoryl-β-D-ribose oxidase (DprE1) and decaprenylphosphoryl-β-D-ribose-2-epimerase (DprE2) catalyze epimerization of decaprenylphosporyl ribose (DPR) todecaprenylphosporyl arabinose (DPA) and are critical for the survival of <i>Mtb</i>. Crystal structures of DprE1 so far reported display significant disordered regions and no structural information is known for DprE2. We used homology modeling, protein threading, molecular docking and dynamics studies to investigate the structural and dynamic features of <i>Mtb</i> DprE1 and DprE2 and DprE1-DprE2 complex. A three-dimensional model for DprE2 was generated using the threading approach coupled with <i>ab initio</i> modeling. A 50 ns simulation of DprE1 and DprE2 revealed the overall stability of the structures. Principal Component Analysis (PCA) demonstrated the convergence of sampling in both DprE1 and DprE2. In DprE1, residues in the 269–330 area showed considerable fluctuation in agreement with the regions of disorder observed in the reported crystal structures. In DprE2, large fluctuations were detected in residues 95–113, 146–157, and 197–226. The study combined docking and MD simulation studies to map and characterize the key residues involved in DprE1-DprE2 interaction. A 60 ns MD simulation for DprE1-DprE2 complex was also performed. Analysis of data revealed that the docked complex is stabilized by H-bonding, hydrophobic and ionic interactions. The key residues of DprE1 involved in DprE1-DprE2 interactions belong to the disordered region. We also examined the docked complex of DprE1-BTZ043 to investigate the binding pocket of DprE1 and its interactions with the inhibitor BTZ043. In summary, we hypothesize that DprE1-DprE2 interaction is crucial for the synthesis of DPA and DprE1-DprE2 complex may be a new therapeutic target amenable to pharmacological validation. The findings have important implications in tuberculosis (TB) drug discovery and will facilitate drug development efforts against TB.</p></div

Cross-Correlated motions of unbound and bound DprE1 and DprE2.

<p>(a) The correlations for the unbound DprE1 are given in upper left triangle and correlations for bound DprE1 are given in the lower right triangle. (b) The correlations for the unbound DprE2 are given in upper left triangle and correlations for bound DprE2 are given in the lower right triangle. Positive correlations are indicated in red and negative correlations in blue.</p

Dual responsive chemosensors for anions : the combination of fluorescent PET (Photoinduced Electron Transfer) and colorimetric chemosensors in a single molecule

The design and synthesis of two novel fluorescent PET anion sensors is described, based on the principle of &lsquo;fluorophore-spacer-(anion)receptor&rsquo;. The sensors 1 and 2 employ simple diaromatic thioureas as anion receptors, and the fluorophore is a naphthalimide moiety that absorbs in the visible part of the spectrum and emits in the green. Upon recognition of anions such as F&minus; and AcO&minus; in DMSO, the fluorescence emission of 1 and 2 was &lsquo;switched off&rsquo;, with no significant changes in the UV&ndash;vis spectra. This recognition shows a 1:1 binding between the receptor and the anions. In the case of F&minus;, further additions of the anion, gave rise to large changes in the UV&ndash;vis spectra, where the &lambda;max at 455 nm was shifted to 550 nm. These changes are thought to be due to the deprotonation of the 4-amino moiety of the naphthalimide fluorophore. This was in fact found to be the case, using simple naphthalimide derivatives such as 6. Sensors 1 and 2 can thus display dual sensing action; where at low concentrations, the fluorescence emission is quenched, and at higher concentrations the absorption spectra are modulated.<br /