Cu(I)-catalyzed "click-chemistry". Design of a chemical photomultiplier. Target-guided synthesis of bidentate metal-complex receptors (2008)

A simple catalytic system, which is able to translate a light signal into a chemical output with amplified response, was developed. The overall amplification factor is 15 after a reaction time of 20 min. Although its fidelity in terms of sensitivity and amplification is far from biological models, the results show that coupling optical and chemical processes allows for the processing and amplification of information. New fluorescent dyes containing alkyne- and azide-groups were synthesized. These compunds allow modular connection to a lot of substrates, such as labelling of proteins. Novel receptor building blocks for different binding motifs have been synthesized. Zn(II)-cyclen complexes for imide groups, bis-Zn(II)-cyclen complexes for phosphate groups, Zn(II) and Cu(II) nitrilo-triacetic acid complexes for N-terminal histidines, guanidines for carboxylic acids and a Zn(II)-porphyrin for non-polar regions on a protein surface. All compounds bear either an azide or an alkyne function and therefore can be connected in any combination using the Cu(I)-catalyzed cycloaddition. A biological relevant pentapeptide, with a histidine and a carboxyl-group as possible binding motifs, has been used as target in a kinetically controlled target guided reaction. In order to identify a bidentate receptor with a high binding constant for the target, combinations of Zn(II)- and Cu(II)-NTA complexes and guanidines have been tested in this reaction. Hit-detection was performed in situ with MS-spectroscopy. Results revealed one possible high affinity receptor out of 12 combinations. Six bidentate receptors have been synthesized and their binding interactions with the peptide target have been investigated with NMR- MS- and fluorescence-spectroscopy. All methods confirmed binding interactions. Results of NMR-investigations allowed a prediction of the structural relations of the formed complex between target and receptor. This prediction was confirmed by Spartan-calculations. Further, the association constant of this complex was determined with fluorescence spectroscopy to be in the micro molar range. However, a definite determination, as well as a comparison of the association constants of these six receptors was not possible. Fluorescent labelled N-terminal histidines, with a dansyl- or a fluorescein group have been synthesized. The binding interactions between the histidine-moiety and heavy metal cations as well as Cu(II)-NTA complexes has been investigated with fluorescence spectroscopy. Results revealed high association constants (> 1011 L2/mol2) of these indicators towards Cu2+ and Cu(II)-NTA complexes. Therefore, these indicators can be used in very sensitive detection assays for Cu(II) and corresponding complexes. Using the Cu(I)-catalyzed cycloaddition, several bidentate receptor molecules with different binding sites have been synthesized. Combinations of receptor building blocks were: bis-Zn(II)-cyclen-complexes with Zn(II)-NTA-complexes or guanidine compounds and further a Cu(II)-NTA- with a Zn(II)-porphyrin complex. The binding interactions between the bis-Zn(II)-cyclen compounds and phosphorylated peptides were investigated with a fluorescence polarisation assay. Results confirmed a slight increase of the binding affinity (factor 2) compared to a monodentate bis-Zn(II)-cyclen reference compound. The binding interactions between the Zn(II)-porphyrin-Cu(II)-NTA complex and amino acids, a pentapeptide and two His-containing proteins (HEL and Myo) were investigated with fluorescence titrations. Results showed a specific binding of histidine as well as of peptides with an N-terminal His. Binding affinities were in the micromolar range. The binding affinities of the porphyrine receptor towards His-containing proteins revealed to be also in the micromolar range, but unspecific. New 1,1�-disubstituted [4,4�]-bitriazole compounds have been synthesized using the Cu(I)-catalyzed cycloaddition between 1,4-butadiyne and various alkyl- and aryl-triazoles. Ru(II)-complexes of the obtained ligands have been synthesized and their characteristics in the UV- and fluorescence spectrum have been determined.