Pyrene Excimer Research Articles

Multi‐Colour Electroluminescence of Dendronic Antennae Containing Pyrenes as Light Harvesters

Dendronic antennae systems containing pyrene units as energy donors and a styrylpyridinium derivative as energy acceptor show efficient energy transfer from the green-emitting pyrene excimer to the red-emitting acceptor. For the third dendron generation the effective screening of the pyrene units on the acceptor provides thin films showing bright red emission. Single-layer light-emitting diodes prepared by properly balancing the dendrons and donor units concentration in polyvinylcarbazole show electroluminescence from the blue, green and red components of the monomeric donor, the donor excimer and the acceptor when excitons are generated in the polymer and subsequently transferred to the molecules by resonant energy transfer.

Syntheses and fluorescence of RNA conjugates having pyrene-modified adenosine and nitrobenzene-modified uridine base pairs

Abstract We prepared RNA duplexes possessing 2′- O -(1-pyrenylmethyl)adenosine and 5-(4-nitrophenyl)uridine base pairs. In the duplexes, pyrene serves as a photo-excitable electron donor and 5-(4-nitrophenyl)uridine acts as an electron acceptor. The donor–acceptor-modified RNA duplexes showed very weak fluorescence originating from the pyrene monomer and excimer emissions, which occur due to electron transfer from the excited pyrene to the nitrobenzene acceptor.

Zinc-cyclen coordination to UTP, TTP or pyrophosphate induces pyrene excimer emission

Pyrene labelled Zn(2+)-cyclen 1 and bis-Zn(2+)-bis-cyclen 2 complexes were synthesized. The reversible coordination at physiological pH of Zn(2+)-cyclens to phosphate anions and to imide moieties, as present in thymine and uracil nucleotides, is well known. In the presence of analytes bearing a phosphate and an imide or two phosphate groups the formation of a ternary complex consisting of two pyrene-labelled metal complexes and the analyte molecule, is observed. The close proximity of the pyrene labels in the complex induces pyrene excimer emission, which is observable by the unarmed eye. By this, the presence of UMP, UDP, UTP and TTP in buffered aqueous solution is signalled, while other nucleotides are not able to induce excimer emission. In the same way, Zn(2+)-cyclen-pyrene acts as luminescent chemosensor for PP(i) and fructose-1,6-bisphosphate in aqueous buffer.

Biochemical Definition of ‘harsh’ Vs. ‘mild’ Detergents For Membrane Protein Solubilization

Selection of a solubilizing detergent for membrane proteins is typically based on its ability to maintain the native structure and/or function of the molecule of interest. Descriptors of detergents as ‘harsh’ or ‘mild’ in terms of their propensity to denature membrane protein structures may act as a qualitative guide to this process, but the basis of the variable effects of detergents on native membrane protein folds is not yet fully explained. Previous work by our group suggested that the ability of the ‘harsh’ detergent sodium dodecylsulfate (SDS) to denature a series of wild-type and mutant model helical membrane-soluble ‘hairpin’ (helix-loop-helix) proteins depends on their level of detergent binding, as manifested by significant variability in their electrophoretic mobilities on SDS-PAGE, and in circular dichroism (CD) spectra and hydrodynamic radii [Rath, Nadeau et al., PNAS USA 106, 1760-65 (2009)]. Here we have extended this work to the characterization of a corresponding library of hairpin proteins in sodium perfluorooctanoate (SPFO), a surfactant considered to be non-denaturing. The ‘hairpin’ library has been developed from helix-loop-helix constructs from transmembrane segments 3 and 4 from the cystic fibrosis transmembrane conductance regulator (CFTR). We find that SPFO-solubilized ‘hairpins’ exhibit significantly less variability vs. their SDS-solubilized counterparts in their electrophoretic mobilities, CD spectra, and hydrodynamic radii. In addition, SPFO favours more stable helical structure while binding hairpins in larger amounts than does SDS across hairpin sequences. Pyrene excimer fluorescence data indicate that SPFO additionally retains more tertiary contacts vs. SDS for all hairpin sequences tested. Solubilization in SPFO therefore appears to favour protein-protein over lipid-protein interactions. Our overall results imply that the ‘harshness’ of a detergent is proportional to its protein sequence specificity upon binding, and consequent tendency not to disrupt intra-protein contacts during micelle formation.

Nucleic acid G-quadruplex based label-free fluorescence turn-on potassium selective sensing

We report a label-free fluorescence turn-on approach for the selective sensing of potassium. A properly selected G-rich oligonucleotide (oligo-Y) folded into stable quadruplex structure when mixed with potassium in an aqueous solution. Single-stranded nucleic acid specific nuclease was subsequently added. Since an oligonucleotide in quadruplex structure is markedly more resistant to nuclease digestion than in its random coil conformation, oligo-Y digestion by nuclease was considerably slow. On the other hand, oligo-Y mixed with other common mono- or divalent ions was completely digested in 5 min under our experimental conditions because no quadruplex or less stable quadruplex was formed. Oligo-Y in potassium was subsequently mixed with a positively charged pyrene probe. Electrostatic interactions between oligo-Y (a polyanion) and the probe induced aggregation of the probe, which in turn induced strong pyrene excimer emission. The intensity of the induced excimer emission was directly proportional to the amount of potassium added. Our method shows good sensitivity, and good selectivity against other common interference ions.

Invader LNA: Efficient targeting of short double stranded DNA

Despite progress with triplex-forming oligonucleotides or helix-invading peptide nucleic acids (PNAs), there remains a need for probes facilitating sequence-unrestricted targeting of double stranded DNA (dsDNA) at physiologically relevant conditions. Invader LNA probes, i.e., DNA duplexes with "+1 interstrand zipper arrangements" of intercalator-functionalized 2'-amino-alpha-l-LNA monomers, are demonstrated herein to recognize short mixed sequence dsDNA targets. This approach, like pseudo-complementary PNA (pcPNA), relies on relative differences in stability between probe duplexes and the corresponding probe:target duplexes for generation of a favourable thermodynamic gradient. Unlike pcPNA, Invader LNA probes take advantage of the "nearest neighbour exclusion principle", i.e., intercalating units of Invader LNA monomers are poorly accommodated in probe duplexes but extraordinarily well tolerated in probe-target duplexes (DeltaT(m)/modification up to +11.5 degrees C). Recognition of isosequential dsDNA-targets occurs: a) at experimental temperatures much lower than the thermal denaturation temperatures (T(m)'s) of Invader LNAs or dsDNA-targets, b) at a wide range of ionic strengths, and c) with good mismatch discrimination. Recognition of dsDNA is monitored in real-time using inherent pyrene-pyrene excimer signals of Invader LNA probes, which provides insights into reaction kinetics and enables rational design of probes. These properties render Invader LNAs as promising probes for biomedical applications entailing sequence-unrestricted recognition of dsDNA.

A unique protein labeling system based on melittin and the non-covalent binding-induced pyrene excimer

We report a unique protein labeling system based on melittin and a pyrene derivative (1). The specific region of the C-terminal in melittin efficiently induced the formation of the pyrene eximer, which can be used as a tag to target proteins and for further detection.

OFF–ON Fluorescent Detection of Thymidine Nucleotides by a Zinc(II)–Cyclen Complex Bearing Two Diagonal Pyrenes

A fluorescent chemosensor consisting of a ZnII–cyclen complex bearing two diagonal pyrenes (see picture) has been developed that has a high affinity for thymidine nucleotides, and exhibits a much greater enhancement in pyrene excimer emission for thymidine triphosphate, -diphosphate and -monophosphate than for the other DNA bases and a variety of nucleotides.

Excited states of pyrene excimer observed by photodissociation spectroscopy in a supersonic jet

The absorption spectrum of jet-cooled pyrene excimer was measured using photodissociation spectroscopy. Broad absorption bands were observed in the near-IR and visible regions, which were assigned to the B 2u − ← B 3g − and B 2u + ← B 3g − transitions of the excimer, respectively. Excitation of these bands results in rapid dissociation of the excimer into monomer fragments, one of which is an electronically excited monomer in the S 2 state. The formation mechanism of the pyrene excimer from the vdW dimer is also discussed.

Pyrene Excimer Nucleic Acid Probes for Biomolecule Signaling

Pyrene excimer nucleic acid probes are novel molecular probes that integrate the unique optical properties of pyrene excimer with excellent recognition capability of nucleic acids. Pyrene has the property of forming excimer that has a large Stokes shift and long fluorescence lifetime. Nucleic acid probes, like aptamers and molecular beacons, can directly bind to target molecules with a high selectivity and affinity. Possessing these outstanding characteristics of pyrene excimer and nucleic acid probes, pyrene excimer nucleic acid probes can be effectively used in detection of biomolecules, study of protein-DNA interactions, and investigation of enzymatic processes in complex biological systems. In this review, we will discuss the optical properties of pyrene excimer and summarize recent progresses on pyrene excimer signaling nucleic acid probes for the signaling of biomolecules and their interactions in complex biological samples.

Unique Sandwich Stacking of Pyrene-Adenine-Pyrene for Selective and Ratiometric Fluorescent Sensing of ATP at Physiological pH

A pincer-like benzene-bridged sensor 1 with a pyrene excimer as a signal source and imidazolium as a phosphate anion receptor was synthesized and investigated for ATP sensing. A unique switch of excimer vs monomer pyrene fluorescence of 1 is observed in the presence of ATP due to the charcteristic sandwich pi-pi stacking of pyrene-adenine-pyrene. On the other hand, four other bases of nucleoside triphosphates such as GTP, CTP, UTP, and TTP can interact only from the outside with the already stabilized stacked pyrene-pyrene dimer of 1, resulting in excimer fluorescence quenching. The fluorescent intensity ratio of monomer-to-excimer for 1 upon binding with ATP (I(375)/I(487)) is much larger than that upon binding with ADP and AMP. This difference is large enough to discriminate ATP from ADP and AMP. As one of the biological applications, sensor 1 is successfully applied to the ATP staining experiments. Sensor 1 is also applied to monitor the hydrolysis of ATP and ADP by apyrase. The results indicate that 1 is a useful fluorescent sensor for investigations of ATP-relevant biological processes.

Pyrene-labeled deoxyguanosine as a fluorescence sensor to discriminate single and double stranded DNA structures: Design of ends free molecular beacons

A novel fluorescent DNA probe containing pyrene-labeled C8 alkylamino-substituted 2′-deoxyguanosine was designed in order to discriminate single stranded and double stranded regions in DNA. This fluorescent sensor was used for the design of practically useful 3′- and 5′-ends free self-quenched molecular beacon (MB). Unique MB detectable by pyrene excimer fluorescence was also demonstrated.

A Light‐Driven Supramolecular Optical Switch

Switched on: The fluorescence emission of a DNA-based optical switch can be reversibly switched from blue to green using modulating light of different wavelengths (see picture). The device is based on the hybridization of two pyrene-functionalized DNA strands in the presence of a photochromic azobenzene-containing effector ligand. Hybridization occurs only with the cis-configured ligand, and results in pyrene excimer fluorescence.

Quantitative analysis of the cross-linked structure of microgels using fluorescent probes

Properties of polymeric microgels are influenced by the internal polymer cross-linked structure, but tools to quantitatively analyze this internal structure are limited. With the finding that polymer networks alter the diffusivity and subsequent excimer formation of pyrene, this study used the ratio between pyrene excimer and monomer emission to determine the number of cross-links (N) and average pore size (ξ) in poly(ethylene glycol) diacrylate (PEGDA) microgels. A calibration curve to relate pyrene emission to N and ξ in PEGDA hydrogels was prepared and used to calculate N and ξ in PEGDA microgels. The pyrene emission indicated that PEGDA microgels had a higher cross-linking density and a smaller average pore size when compared with bulk cross-linked hydrogels of the same PEGDA concentration. The analytical method demonstrated in this study may be useful for fine-tuning polymeric microgel properties for a broad array of applications.

Probing the microenvironment of surface-attached pyrene formed by a thermo-responsive oligomer

We have constructed a novel molecular assembly attached to quartz, oxidized silicon and indium-doped tin oxide coated substrates, where a tethered pyrene derivative is co-immobilized with oligo- N-isopropyl acrylamide (oligo-NIPAM). The addition of tethered oligo-NIPAM to the adlayer creates two different, temperature-dependent microenvironments for the surface-bound pyrene. X-ray photoelectron spectroscopy (XPS) and ellipsometry measurements demonstrate the covalent attachment of both oligo-NIPAM and pyrene in our adlayers. Contact angle results confirm the thermo-responsive nature of the oligo-NIPAM on the substrate surface. Steady-state fluorescence data show that the presence of oligo-NIPAM moieties reduces the extent of pyrene excimer formation and provides different environments for the chromophore at temperatures above and below the phase transition. Fluorescence lifetime decay data on surface-bound pyrene are biexponential, consistent with multiple local environments, regardless of whether tethered oligo-NIPAM is present or not. Quenching studies reveal that we can manipulate the sensing properties of this new film simply by adjusting the conformations of oligo-NIPAM.

Nucleic Acid-Induced Aggregation and Pyrene Excimer Formation

Nucleic acid was found to induce the aggregation of the positively charged pyrene probe (compound 1); as a result, strong pyrene excimer emission was observed. The intensity of the excimer emission was dependent on the concentration of the pyrene probe and the oligonucleotide length, sequence, and concentration. These results suggest a new strategy for label-free nucleic acid-based biosensing applications.

Design of extremely facile 3'- and 5'- ends free molecular beacons using C8 alkylamino substituted 2'-deoxyguanosine

A novel fluorescent oligonucleotide probe containing pyrene-labeled C8 alkylamino-substituted 2'-deoxyguanosine and a practically useful 3'- and 5'- ends-free self-quenched molecular beacons (MB) were designed. A unique MB detectable by pyrene excimer fluorescence was also demonstrated.

Lithography Based on Memory Effects Resulting from Photoinduced Self-Assembly of Pyrene Dimers in Thin Polymer Films

Pulsed laser irradiation of pyrene in thin films through lithographic masks leads to blue fluorescent images characteristic of pyrene excimer emission. The images are stable at room temperature and are readily observable using fluorescence microscopy. For a constant energy dose, the images are stronger and better resolved at the shorter laser wavelengths and cannot be recorded with continuous wavelength (CW) light sources. Images are easier to obtain with high Tg films, while polymers with Tg below room temperature image only when the laser also causes polymer cross-linking. It is proposed that laser excitation generates excited singlet pyrene and simultaneously causes a temperature change that facilitates diffusion and thus excimer formation. Following excited state (monomer and excimer) decay, the polymer matrix cools down and prevents the separation of the pyrene pairs, that are preserved by the rigid polymer matrix. Some of the images can be thermally erased by annealing above Tg. The method may find applications in imaging and information storage. © 2009 American Chemical Society.

Stage-specific changes in membrane microviscosity in Misgurnus fossilis embryos

Natural and probe fluorescence as well as membrane microviscosity was studied in eggs and embryos of Misgurnus fossilis by fluorescence microscopy. The lateral mobility of the probe (pyrene) increased in loach embryos from early to late blastula, which indicates a decrease in plasma membrane microviscosity. At the later stage of mid-gastrula, the microviscosity remained largely invariant. Considering that the embryo exposure to different temperatures changes the quantum yield of fluorescence and the degree of pyrene excimerization, one can gain information about both the temperature-induced structural changes and changes in membrane microviscosity in the embryos. Natural and probe fluorescence of embryonic membranes is proposed as at tool to study morphogenetic mechanisms.

Solvent dependence of pyrene monomer and excimer fluorescence quenching by heavy atoms

Two independent methods have been used for the determination of pyrene monomer (M*) and excimer (D*) quenching rate parameters in eight common solvents ranging in viscosities between 0.3 and 5.9 cP. Heavy atoms (CCl4) were used as quenchers. The discrepancies between the values of the quenching rate parameters reported in the two methods have been explained as due to formation of complexes between the pyrene molecule and CCl4 molecules as a result of using a more intense source, which might lead to larger values of the quenching rate parameters. Assuming that the quenching processes are diffusion-controlled ones, the probabilities of the effective collisions between M* or D* and CCl4 (PP or PP, respectively) were then calculated. These values were found to depend on the nature of the solvents used and in particular on their viscosities (η). At low values of η, the average values of PP and PP are approximately the same for each solvent used. For intermediate values of η, PP becomes much larger than PP. Finally for relatively large values of η, PP becomes much smaller than PP.