Monitoring interfacial bioelectrochemistry using a FRET switch

Generation of functionally active biomolecular monolayers is important in both analytical science and biophysical analyses. Our ability to monitor the redox-active state of immobilized proteins or enzymes at a molecular level, from which stochastic and surface-induced variations would be apparent, is impeded by comparatively slow electron-transfer kinetics and associated signal:noise difficulties. We demonstrate herein that by covalently tethering an appropriate dye to the copper protein azurin a highly oxidation-state-sensitive FRET process can be established which enables redox switching to be optically monitored at protein levels down to the zeptomolar limit. The surface-potential-induced cycling of emission enables the redox potential of clusters of a few hundred molecules to be determined.     

Transport of noble gases in poly(methyl acrylate)

The permeability, diffusivity, and solubility of He, Ne, Ar, and Kr were determined in poly(methyl acrylate) (PMA), in a temperature range encompassing the glass transition temperature, T_g. Activation energies for diffusion, E_D, were higher above Tg than below Tg for all four penetrants in PMA and in the structural isomer of PMA, poly(vinyl acetate) (PVA). For all penetrants studied, the Tg, the magnitude of the enthalpy of mixing (ΔH_m), as well as the E_D were all larger for PVA than for PMA. These differences were attributed to the stronger dipole-dipole interactions possible in PVA where the dipolar carbonyl group is separated from the chain backbone by an oxygen atom.

The carbonyl group in PMA is immediately adjacent and presumably sterically hindered by the chain backbone which suggests that PMA might be a stiffer molecule than PVA. Entropy considerations suggest that molecularly stiff polymers should be associated with small values of v Δα, ΔE_D, ΔE_D, ΔH_p, and (β), where v is the specific volume, Δα is the change in thermal expansivity about T_g, ΔH_p is the change in activation energy for diffusion about Tg, ΔE_D is the enthalpy of polymerization, and (β) is the logarithmic bulk relaxation rate constant of specific volume below T_g. These predictions appear to be satisfied for the systems of argon in PVC, PMA, PVA, and poly(ethyl methacrylate) (PEMA). The data suggest that PMA is a stiffer molecule than PVA. The weaker intermolecular forces of attraction and more hindered molecular rotations of PMA are consistent with a priori considerations of steric effects which have their origin in the fundamental structure of the PMA and PVA molecules.

The fraction of volume, φ, not occupied by polymer, estimated by several independent means for PMA at T_g, appears to be greater than that of PVA. Each of these two comparisons suggests that the molecular packing of PMA is more dense than the molecular packing of PVA.

The observed slow decrease in specific volume for PMA below Tg suggests that there could be regions throughout the polymer below Tg where the density is less than the equilibrium value. These predictions are consistent with the analyses of penetrant solubility below Tg which suggest that the molal volumes of the penetrants below Tg are larger than those above Tg. Also, analyses of the solubility data suggest that the enthalpies of mixing the noble gases in PMA tend to be exothermic below Tg. This suggests that below Tg, specific interactions occur between PMA and noble gas penetrants. The interactions are apparently manifested by decreases in free energy which are larger than expected for inert penetrants. The solubility and volumetric data appear to be consistent with the hypothesis that the polymer below Tg contains expanded, high-energy regions which preferentially absorb the diffusing penetrants.     

Packing as a probe of structure in alkaline earth glass systems

Packing is an intrinsic property of glass, defined as the ratio of ionic volume to molar volume, and is a useful parameter for analyzing structural changes with composition. Alkali based glasses show two trends in packing, one dominated by the oxygen covalent network for the small ions, Li and Na, and one ionically dominated by the metal cations for the large, K, Rb and Cs cases [S. Giri, C. Gaebler, J. Helmus, M. Affatigato, S.A. Feller, J. Non-Cryst. Solids 347 (2004) 87]. We have found that alkaline earth glasses do not display these behaviors, and in this paper we determined the packing fractions of these glasses and compared them with the alkali case. Further, we considered the structural implications of the packing trends.     

Enthalpies of reaction of bis(triphenylphosphine)(trans-stilbene)platinum(0) with diphenylcyclopropenone and benzocyclobutenedione

Enthalpies, ΔH(1) ?94.8 ± 6.0 and ΔH(6) ?57.1 ± 5.1 kJ mol^?1, of the following reactions have been measured calorimetrically [Pt(trans-stilbene)(PPh₃)₂](s) + dpcp(g) → (PPh₃)₂Pt(dpcb)(s) + trans-stilbene(g) (1) [Pt(trans-stilbene)(PPh₃)₂](s) + bcbd(g) → (PPh₃)₂Pt(bcpd)(s) + trans-stilbene (g) (6) where dpcp is diphenylcyclopropenone, (PPh₃)₂Pt(dpcb) is (1,1-bistriphenylphosphine)platinadiphenylcyclobutenone, (PPh₃)₂$\text{PtC(Ph)C(Ph)C}$O, bcbd is benzocyclobutene-1,2-dione and (PPh₃)₂Pt(bcpd) is (1,1-bistriphenylphosphine)platinabenzocyclopentanedione,

. It is concluded that the five-membered platinacyclo ring system in (PPh₃)₂Pt(dpcb) is not heavily strained.     

A combinatorial method for solution-phase synthesis of labeled bivalent beta-turn mimics

Piperidine-functionalized, 1,4-disubstituted-1,2,3-triazoles of generic structure 1 were conceived as "minimalist" mimics of peptidic beta-turn structures. Key features of these molecules include (i) the possibility of incorporating amino acid side chains corresponding to many of the protein amino acids; (ii) a close correspondence of separations of these side chains to i + 1 to i + 2 residues in turns; (iii) facile adjustment of the side-chain vectors on docking while only influencing two critical degrees of freedom; and (iv) some electrostatic polarity. Fifteen monomers of this type were made via copper-mediated cycloaddition reactions. Solution-phase methodologies were devised to assemble these monomers into bivalent compounds in high purity states (typically >85%) so that they could be used in first-pass biological assays without further purification. The skeleton for forming these bivalent compounds is triazine-based. There is a third site which allowed for introduction of a fluorescent label (library of compounds 2) or an alkyne-functionalized triethylene glycol chain (library of compounds 3) included to promote water-solubility and to allow incorporation of probes via copper-mediated cycloaddition reactions. In the event, two 135-membered libraries were prepared, one consisting of compounds 2 and the other of 3. No protecting groups or coupling agents were required; these attributes of the method were important to allow most of the products to be obtained in over 85% purities. The fluorescein-tagged library of compounds 2 was screened in a fluorescence-activated cell sorting (FACS) assay using cells transfected to overexpress one of the following neurotrophin receptors: TrkA, TrkC, and p75. Preliminary findings indicate four compounds 2gm, 2gn, 2gi, and 2gj bound the TrkA receptor selectively; all of these contain a threonine-lysine turn mimic. Thus, a pharmacological probe for the TrkA receptor has been developed.     

Heterovalent selectivity and the combinatorial advantage

Libraries of monovalent compounds can be reacted with each other to give libraries of bivalent ones. If those reactions are efficient, and if the products do not need to be purified, large numbers of bivalent compounds can be produced rapidly, and one might say there is a "combinatorial advantage" to doing so. However, selective formation of heterobivalent products must be possible otherwise statistical mixtures will form. This tutorial review describes methods that will give heterobivalent compounds almost exclusively. Although there are relatively few methods that will give that desired selectivity, such methods are becoming increasingly important as the potential applications of bi- and multivalent compounds emerge.     

Syntheses and properties of water-soluble Nile Red derivatives

Nile Red (compound A) fluoresces at about 530 nm with good quantum yields in apolar solvents. In more polar ones its fluorescence emission shows a dramatic, and potentially useful, shift to about 640 nm, but its quantum yield is significantly reduced. Further, Nile Red has a very poor solubility in aqueous media. The hypothesis tested in this paper is that Nile Red derivatives that incorporate water-solubilizing groups will tend to fluoresce with good quantum yields in aqueous media, and in the more useful wavelength range around 640 nm. Thus three Nile Red derivatives, 1-3, were prepared. Compound 1 had three hydroxyl groups more than Nile Red, but was surprisingly insoluble in aqueous media. However, the dicarboxylic acid 2 and carboxylic/sulfonic acid derivative 3 showed excellent water solubilities. Spectral data for 2 and 3 showed that they do indeed fluoresce with good quantum yields in the 640 nm region in aqueous media. These properties of compounds 2 and 3 might be useful in the development of fluorescent probes for biotechnology.