pyrophosphate and 2-2--dipicolylamine

Phosphate oxoanions and phosphorylated biomolecules (such as nucleotides, lipids, and proteins) play key roles in a wide range of biological processes. The ability to selectively detect these ions in the presence of each other has numerous applications in biochemistry and biomedicine. However, receptors and sensors that can discriminate between polyphosphate species with high selectivity and in biologically relevant conditions are rare. In this Account, we show how peptides (both cyclic and linear) can be used to position two zinc(II)dipicolylamine [Zn(II)DPA] binding sites in an appropriate arrangement to provide selective binding of pyrophosphate (PPi) in the presence of other polyphosphate species, including ATP, and in complex media such as cell growth buffer. The use of peptide scaffolds to position the Zn(II)DPA anion binding sites allowed the synthesis of small receptor libraries in which the arrangement of the two binding sites could be subtly altered to evaluate the factors affecting both binding selectivity and affinity for PPi. We altered a number of structural elements including peptide length, cyclic peptide ring size, amino acid composition, the positioning of the binding sites with respect to one another, and the relative stereochemistry of the peptides. Backbone modified cyclic peptides based on the Lissoclinum class of natural products were initially employed to provide an added degree of preorganization to the receptors, although it was subsequently found that short, flexible bis[Zn(II)DPA]-functionalized linear peptides are also effective scaffolds for selective pyrophosphate recognition. The peptidic receptors were successfully employed for the detection of PPi in aqueous media by indicator displacement assays using both colorimetric and fluorescent indicators, with the best compounds able to bind to PPi selectively in both cell growth media and artificial urine and also allow the accurate determination of PPi concentrations in physiologically relevant ranges (micromolar concentrations) in these complex media. Improved pyrophosphate selectivity was observed upon increasing the complexity of the media from HEPES buffer to cell growth media (Krebs saline). Pyrophosphate sensors in which a fluorescent indicator was covalently attached to either a linear or cyclic peptide scaffold through a flexible linker were then constructed. When the Zn(II)DPA binding sites and the indicator were suitably placed with respect to one another on the pepti

Topics: Amines; Binding Sites; Diphosphates; Fluorescent Dyes; Peptides; Picolinic Acids; Water; Zinc

The label-free detection of pyrophosphate (PPi) anions with a nanofluidic sensing device based on asymmetric nanopores is demonstrated. The pore surface is functionalized with zinc complexes based on two di(2-picolyl)amine [bis(DPA)] moieties using carbodiimide coupling chemistry. The complexation of zinc (Zn(2+) ) ion is achieved by exposing the modified pore to a solution of zinc chloride to form bis(Zn(2+) -DPA) complexes. The chemical functionalization is demonstrated by recording the changes in the observed current-voltage (I-V) curves before and after pore modification. The bis(Zn(2+) -DPA) complexes on the pore walls serve as recognition sites for pyrophosphate anion. The experimental results show that the proposed nanofluidic sensor has the ability to sense picomolar concentrations of PPi anion in the surrounding environment. On the contrary, it does not respond to other phosphate anions, including monohydrogen phosphate, dihydrogen phosphate, adenosine monophosphate, adenosine diphosphate, and adenosine triphosphate. The experimental results are described theoretically by using a model based on the Poisson-Nernst-Planck equations.

Topics: Amines; Diphosphates; Nanopores; Picolinic Acids; Staining and Labeling

A microarray-chip assay system for the fluorescence detection of phosphate-containing analytes in aqueous media has been constructed from stimuli-responsive polymerized poly(diacetylene)-liposomes for the first time. Proper combination of the liposome components (Zn(II)-dipicolylamine for phosphate binding and an amine-terminated component for anchoring the liposome onto an aldehyde-derivatized glass plate), has led to a microarray chip that selectively detects pyrophosphate, an important biomarker, over competing anions, such as phosphate and adenosine triphosphate, with nanomolar sensitivity. The chip-based assay shows advantages, such as high specificity and sensitivity, over solution-based assays that use the same liposomes, and over known homogeneous molecular sensing systems.

Topics: Amines; Diphosphates; Fluorescence; Liposomes; Microarray Analysis; Microscopy, Electron, Scanning; Models, Molecular; Picolinic Acids; Polyacetylene Polymer; Polymers; Polyynes; Zinc

A new zinc(II) complex with a two-dipicolylamine-substituted 1,8-naphthalimide for recognition of pyrophosphate with ratiometrical fluorescence changes in aqueous solution has been synthesized and characterized. Its biological application to monitor the intracellular pyrophosphate (PPi) was successfully demonstrated by the observation that the fluorescence of 1 was enhanced by the presence of the Zn(2+) ion and was quenched by addition of PPi.

Topics: Amines; Animals; Cell Line; Diphosphates; Fluorescent Dyes; Mice; Models, Molecular; Molecular Structure; Naphthalimides; Picolinic Acids; Zinc

Mononuclear Zn(II)-DPA and Cu(II)-DPA complexes crafted on 2-hydroxy-6-cyanonaphthalene fluorophore selectively recognize PPi over ATP and other anions including inorganic phosphates in aqueous medium, showing turn-on type fluorescence enhancements. Coordination of a hydroxyl group of the fluorophore, directly or in alkoxy form, to the central metal ion is crucial for the sensing processes. Both the complexes elicit a fluorescence increase in a time-dependent fashion.

Topics: Amines; Anions; Copper; Diphosphates; Fluorescence; Fluorescent Dyes; Molecular Probes; Naphthols; Organometallic Compounds; Phosphates; Picolinic Acids; Solutions; Spectrometry, Fluorescence; Water; Zinc

Recently, significant advances have been made independently in electrogenerated chemiluminescence (ECL) analysis and supramolecular anion sensing. Herein, we demonstrate a new proof of concept for ECL-based pyrophosphate (PPi) sensing, where the emission intensity is changed by electrochemical turn-on. The ECL PPi sensor (1-2Zn) consists of two orthogonally bonded moieties: boron dipyrromethene (ECL reporter) and a phenoxo-bridged bis(Zn(2+)-dipicolylamine) complex (PPi receptor). The presence of PPi is confirmed from the change in the intensity of green ECL generated from the former when PPi is selectively recognized by the latter. During PPi recognition, changes are caused in the electronic states of the receptor, and this stimulates the attenuation of ECL intensity. The electrochemical "on-off" triggering of light emission upon anion binding forms the basis of a new anion sensing strategy. We expect that green-colored ECL sensing would offer an advantage to current ECL analysis.

Topics: Amines; Anions; Boron; Coordination Complexes; Diphosphates; Electrochemical Techniques; Luminescent Measurements; Picolinic Acids; Porphobilinogen; Thermodynamics; Zinc

Two new coumarin derivatives displayed highly selective "Off-On" fluorescence changes with pyrophosphate between various anions including ATP, ADP, AMP and inorganic phosphate in 100% aqueous solution.

Topics: Amines; Clinical Laboratory Techniques; Copper; Coumarins; Crystallography, X-Ray; Diphosphates; Fluorescence; Phosphates; Picolinic Acids