Aaron M. Massari

Substrate binding and protein conformational dynamics measured by 2D-IR vibrational echo spectroscopy

Enzyme structural dynamics play a pivotal role in substrate binding and biological function, but the influence of substrate binding on enzyme dynamics has not been examined on fast time scales. In this work, picosecond dynamics of horseradish peroxidase (HRP) isoenzyme C in the free form and when ligated to a variety of small organic molecule substrates is studied by using 2D-IR vibrational echo spectroscopy. Carbon monoxide bound at the heme active site of HRP serves as a spectroscopic marker that is sensitive to the structural dynamics of the protein. In the free form, HRP assumes two distinct spectroscopic conformations that undergo fluctuations on a tens-of-picoseconds time scale. After substrate binding, HRP is locked into a single conformation that exhibits reduced amplitudes and slower time-scale structural dynamics. The decrease in carbon monoxide frequency fluctuations is attributed to reduced dynamic freedom of the distal histidine and the distal arginine, which are key residues in modulating substrate binding affinity. It is suggested that dynamic quenching caused by substrate binding can cause the protein to be locked into a conformation suitable for downstream steps in the enzymatic cycle of HRP.

Neuroglobin dynamics observed with ultrafast 2D-IR vibrational echo spectroscopy

Neuroglobin (Ngb), a protein in the globin family, is found in vertebrate brains. It binds oxygen reversibly. Compared with myoglobin (Mb), the amino acid sequence has limited similarity, but key residues around the heme and the classical globin fold are conserved in Ngb. The CO adduct of Ngb displays two CO absorption bands in the IR spectrum, referred to as N3 (distal histidine in the pocket) and N0 (distal histidine swung out of the pocket), which have absorption spectra that are almost identical with the Mb mutants L29F and H64V, respectively. The Mb mutants mimic the heme pocket structures of the corresponding Ngb conformers. The equilibrium protein dynamics for the CO adduct of Ngb are investigated by using ultrafast 2D-IR vibrational echo spectroscopy by observing the CO vibrations spectral diffusion (2D-IR spectra time dependence) and comparing the results with those for the Mb mutants. Although the heme pocket structure and the CO FTIR peak positions of Ngb are similar to those of the mutant Mb proteins, the 2D-IR results demonstrate that the fast structural fluctuations of Ngb are significantly slower than those of the mutant Mbs. The results may also provide some insights into the nature of the energy landscape in the vicinity of the folded protein free energy minimum.

Organic photovoltaics interdigitated on the molecular scale

Phosphonated porphyrin frameworks that are porous to an electron-accepting perylenediimide are systematically interdigitated with a phosphonated form of the diimide to form bulk heterojunctions. When employed in photovoltaics, these molecularly interlaced heterojunctions provide large junction areas while retaining the phase connectivity of traditional bilayer heterojunctions. Zirconium phosphonate linkages facilitate layer-by-layer chromophore assembly from solution under ambient conditions. The dependence of photovoltaic performance on heterojunction architecture is investigated.

Development and application of patterned conducting polymer thin films as chemoresponsive and electrochemically responsive optical diffraction gratings

Abstract We describe the preparation of two-dimensionally patterned polyaniline (PANI) thin films via microtransfer molding and electropolymerization techniques. This procedure yields reproducible conducting polymer patterns with excellent feature periodicity, making them useful as diffraction gratings. The fabricated polymer gratings were characterized via tapping-mode atomic force microscopy. Spectroelectrochemistry was used to characterize the optical properties associated with various intrinsic PANI redox states. In accordance with the Kramers–Kronig relation for change-in-absorptivity and change-in-index-of-refraction, electrochemically induced changes in refractive index (detected via changes in diffraction efficiency) were observed to coincide with electrochemically-induced changes in the PANI electronic absorption spectrum. In addition, the higher oxidation states of PANI and the associated changes in refractive index proved accessible via chemical oxidation. Beyond the novelty of a chemically-switchable transmission grating, the response of this system points to the possibility of developing diffraction-based chemical sensing schemes.

Ultrathin micropatterned porphyrin films assembled via zirconium phosphonate chemistry

The synthesis of a phosphonic-acid-functionalized porphyrin is presented and a procedure for the reproducible assembly of the porphyrins into thin films on glass or conductive glass surfaces is described. The assembly scheme, which utilizes established zirconium phosphonate (ZrP) chemistry, yields highly oriented films (normal to the surface) of well-defined thicknesses. In the lateral direction (plane parallel to the surface) the porphyrins interact by edge-on-edge contact and are characterized by significant porosity. Electrochemical redox-probe experiments indicate the existence of openings or pores of several angstroms in width in both monolayer and multilayer ZrP porphyrin films. Micropatterned versions of the films, capable of diffracting visible light, have also been prepared and have been used for the direct evaluation of film thicknesses via atomic force microscopy.

Origins of spectral broadening in iodated Vaska's complex in binary solvent mixtures.

Linear absorption spectroscopy of the iridium-bound carbonyl on an iodated adduct of Vaskas complex has shown that the mean vibrational frequency is insensitive to solvation by a broad range of solvents, while the spectral line width changes significantly. The spectral broadening is more significant in chloroform than benzyl alcohol, which is puzzling considering that benzyl alcohol is more polar. In this study, 2D-IR spectroscopy was performed on this vibrational mode to dissect the linear line shape into its homogeneous and inhomogeneous contributions in binary solvent mixtures of either chloroform or benzyl alcohol in d6-benzene. The full frequency-frequency correlation function was determined, including the homogeneous line width and fast spectral diffusion. We find that the frequency fluctuation magnitudes show the most notable changes in chloroform mixtures, while the time constants for spectral diffusion change more dramatically in benzyl alcohol mixtures. Nonetheless, we conclude that the frequency fluctuation magnitudes in both solvent mixtures most clearly explain the differences in their linear line widths. The homogeneous contributions were found to either stay the same or decrease as the more polar solvent was added to d6-benzene, thereby implicating inhomogeneous dynamics as the dominant broadening mechanism.

Solvent-Mediated Vibrational Energy Relaxation from Vaska’s Complex Adducts in Binary Solvent Mixtures

A vibrational pump-probe and FTIR study was performed on two different adducts of Vaskas complex in two different sets of binary solvent mixtures. The carbonyl vibrational mode in the oxygen adduct exhibits solvatochromic shifts of ~10 cm(-1) in either benzyl alcohol or chloroform relative to benzene-d6, whereas this vibration is nearly unchanged for the iodine adduct for the same three solvents. The width and center frequency of the carbonyl stretch for each adduct are compared to its vibrational lifetime in binary mixtures of benzene-d6 with either benzyl alcohol or chloroform. In neat solvents, the trends in line width, frequency, and vibrational lifetime are consistent for the two adducts, but complex relationships emerge when the trends in each property are compared as a function of mixed solvent composition. ν(CO) is more sensitive to the solvation environment around the trans ligand, whereas the line width and lifetime depend on the environment around the CO group itself. The carbonyl frequency and width vary nonlinearly across the two binary solvent series, indicating preferential solvation. In contrast, the vibrational lifetime changes linearly with solvent composition and is correlated with the mole fraction of chloroform but anticorrelated with the mole fraction of benzyl alcohol. The results are explained by differences in the densities of solvent modes that affect intermolecular relaxation of the carbonyl mode.

Experimental evidence for an optical interference model for vibrational sum frequency generation on multilayer organic thin film systems. II. Consideration for higher order terms

The generalized optical interference model for interfacial contributions to vibrational sum frequency generation (VSFG) spectroscopic signals from organic thin film systems is extended to include a description of optical interferences contained in the thin film bulk response. This is based on electric quadrupolar interactions with the input fields and includes a discussion on possible contribution from the electric quadrupolar polarization. VSFG data from the first of this two part report are analyzed and include effects from higher order responses, for both bulk and higher order interfacial terms. The results indicate that although it is capable of capturing many of the data features, the electric dipole treatment is likely not a complete description of the VSFG intensity data from this system. An analysis based on the signs of the resulting response amplitudes is used to deduce the relative magnitude of the electric dipole and higher order interfacial terms. It is found that the buried interface is closer to satisfying the electric dipole approximation, consistent with smaller field gradients due to closer index matching between the organic thin film and substrate relative to air. The procedure outlined in this work allows for the difficult task of deducing a physical picture of average molecular orientation at the buried interface of a multilayer organic thin film system while including higher order effects.

Real-time structural evolution at the interface of an organic transistor during thermal annealing

Polarization multiplexed vibrational sum frequency generation (PM-VSFG) spectroscopy has been used to monitor the interfacial structure of polymer transistor interfaces in situ during thermal annealing treatments. The evolution of the field-effect carrier mobility is tracked simultaneously with the molecular orientation and ordering of poly(3-hexylthiophene) (P3HT) macromolecules on two different surface types. It is shown that fluorocarbon functionalized silica imparts very different molecular arrangements that avoid kinetic trapping during solution casting. In contrast, bare silica surfaces produce kinetically trapped polymer configurations that can be observed by PM-VSFG to reorient with thermal annealing. The interfacial results are compared to bulk structural changes in P3HT thin films as characterized by differential scanning calorimetry and linear spectroscopies. The electrical performances of these films are more closely correlated with interfacial parameters than the bulk properties of the polymer. In contrast with the bulk measurements, the PM-VSFG studies show that molecules at the organic/dielectric interface are actually less ordered after thermal annealing processes that render them with lower carrier mobilities.

Modeling multilayer thin film interference effects in interface-specific coherent nonlinear optical spectroscopies

We develop a generalized model to describe thin film interference in interface-specific nonlinear optical spectroscopies of ideal isotropic stratified systems that enables the separation of this effect from the individual interfacial nonlinear responses. The model utilizes a property of the transfer matrix formalism that allows for simplification of an arbitrary layered system to a single layer with newly defined coefficients of reflection and transmission. In addition to the already well known internal transfer coefficients that relate incident fields to internal fields, we define external transfer coefficients that describe how internally generated fields propagate out of the system. By applying the usual boundary conditions we are able to analytically describe the local and induced fields immediately adjacent to an arbitrary interface, followed by transfer of the generated fields out of the system. The model provides a complete and easily implemented approach to calculating the observables from interface-specific spectroscopies on arbitrary layered thin film systems in a concise way.