Rational design and investigation of nonlinear optical response properties of pyrrolopyrrole aza-BODIPY-based novel push-pull chromophores.

Intramolecular charge transfer (ICT)-based chromophores are highly sought after for designing near-infrared (NIR) absorbing and emitting dyes as well as for designing materials for nonlinear optical (NLO) applications. The properties of these 'push-pull' molecules can easily be modified by varying the electronic donor (D) and acceptor (A) groups as well as the π-conjugation linker. This study presents a methodical approach and employs quantum chemical analysis to explore the relationship between the structural features, electro-optical properties, and the NLO characteristics of molecules with D-π-A framework. The one- and two-photon absorption (2PA), linear polarizability (α), and first hyperpolarizability (ß) of some novel chromophores, consisting of a dimeric aza-Boron Dipyrromethene (aza-BODIPY) analogue, called, pyrrolopyrrole aza-BODIPY (PPAB), serving as the acceptor, have been investigated. The electronic donors used in this study are triphenylamine (TPA) and diphenylamine (DPA), and they are conjugated to the acceptor via thienyl or phenylene π-linkers. Additionally, the Hyper-Rayleigh Scattering (ßHRS), which enables direct estimation of the second-order NLO properties, is calculated for the studied chromophores with 1064 nm excitation in acetonitrile. The ß value shows a significant increase with increasing solvent polarity, indicating that the ICT plays a crucial role in shaping the NLO response of the studied molecules. The enhancement of the 2PA cross-section of the investigated molecules can also be achieved by modulating the combinations of donors and linkers. The results of our study indicate that the novel D-π-A molecules designed in this work demonstrate considerably higher hyperpolarizability values than the standard p-nitroaniline, making them promising candidates for future NLO applications.

Impact of protein-chromophore interaction on the retinal excited state and photocycle of Gloeobacter rhodopsin: role of conserved tryptophan residues.

The function of microbial as well as mammalian retinal proteins (aka rhodopsins) is associated with a photocycle initiated by light excitation of the retinal chromophore of the protein, covalently bound through a protonated Schiff base linkage. Although electrostatics controls chemical reactions of many organic molecules, attempt to understand its role in controlling excited state reactivity of rhodopsins and, thereby, their photocycle is scarce. Here, we investigate the effect of highly conserved tryptophan residues, between which the all-trans retinal chromophore of the protein is sandwiched in microbial rhodopsins, on the charge distribution along the retinal excited state, quantum yield and nature of the light-induced photocycle and absorption properties of Gloeobacter rhodopsin (GR). Replacement of these tryptophan residues by non-aromatic leucine (W222L and W122L) or phenylalanine (W222F) does not significantly affect the absorption maximum of the protein, while all the mutants showed higher sensitivity to photobleaching, compared to wild-type GR. Flash photolysis studies revealed lower quantum yield of trans-cis photoisomerization in W222L as well as W222F mutants relative to wild-type. The photocycle kinetics are also controlled by these tryptophan residues, resulting in altered accumulation and lifetime of the intermediates in the W222L and W222F mutants. We propose that protein-retinal interactions facilitated by conserved tryptophan residues are crucial for achieving high quantum yield of the light-induced retinal isomerization, and affect the thermal retinal re-isomerization to the resting state.

Retinal-Carotenoid Interactions in a Sodium-Ion-Pumping Rhodopsin: Implications on Oligomerization and Thermal Stability.

Microbial rhodopsin (also called retinal protein)-carotenoid conjugates represent a unique class of light-harvesting (LH) complexes, but their specific interactions and LH properties are not completely elucidated as only few rhodopsins are known to bind carotenoids. Here, we report a natural sodium-ion (Na+)-pumping Nonlabens (Donghaeana) dokdonensis rhodopsin (DDR2) binding with a carotenoid salinixanthin (Sal) to form a thermally stable rhodopsin-carotenoid complex. Different spectroscopic studies were employed to monitor the retinal-carotenoid interaction as well as the thermal stability of the protein, while size-exclusion chromatography (SEC) and homology modeling are performed to understand the protein oligomerization process. In analogy with that of another Na+-pumping protein Krokinobacter eikastus rhodopsin 2 (KR2), we propose that DDR2 (studied concentration range: 2 × 10-6 to 4 × 10-5 M) remains mainly as a pentamer at room temperature and neutral pH, while heating above 55 °C partially converted it into a thermally less stable oligomeric form of the protein. This process is affected by both the pH and concentration. At high concentrations (4 × 10-5 to 2 × 10-4 M), the protein adopts a pentamer form reflected in the excitonic circular dichroism (CD) spectrum. In the presence of Sal, the thermal stability of DDR2 is increased significantly, and the pigment is stable even at 85 °C. The results presented could have implications in designing stable rhodopsin-carotenoid antenna complexes.

Conjugation of native membranes via linear oligo-amines.

We report the first observation of an efficient, native membrane conjugation mechanism via positively charged, linear oligo-amines. Clustering of membrane fragments relies on electrostatic interactions between the net negative charge of the membranes and the positively charged, water-soluble mediators. This conjugation principle is demonstrated with two different bacterial membranes in which are embedded either the intrinsic membrane protein (MP) bacteriorhodopsin (bR) or the more recently identified xanthorhodopsin (XR). As determined by their characteristic UV-vis absorption spectra and by circular dichroism, the MPs are not significantly perturbed by the oligo-amines carrying from +3 to +6 positive charges. Light microscopy and scanning electron microscope (SEM) imaging provide direct evidence for membrane conjugation. Process efficiency was found to be correlated with the net charge of the oligo-amine used. Membrane conjugation is accomplished within a wide range of pH values (7-2.5); is reversed by NaCl; and does not require the presence of a precipitant (e.g. PEG) nor Ca2+ ions. Some evidence for bilayer fusion is also observed, but only in the presence of the +6 oligo-amine analog.

Molecular mechanism for thermal denaturation of thermophilic rhodopsin.

Understanding the factors affecting the stability and function of proteins at the molecular level is of fundamental importance. In spite of their use in bioelectronics and optogenetics, factors influencing thermal stability of microbial rhodopsins, a class of photoreceptor protein ubiquitous in nature are not yet well-understood. Here we report on the molecular mechanism for thermal denaturation of microbial retinal proteins, including, a highly thermostable protein, thermophilic rhodopsin (TR). External stimuli-dependent thermal denaturation of TR, the proton pumping rhodopsin of Thermus thermophilus bacterium, and other microbial rhodopsins are spectroscopically studied to decipher the common factors guiding their thermal stability. The thermal denaturation process of the studied proteins is light-catalyzed and the apo-protein is thermally less stable than the corresponding retinal-covalently bound opsin. In addition, changes in structure of the retinal chromophore affect the thermal stability of TR. Our results indicate that the hydrolysis of the retinal protonated Schiff base (PSB) is the rate-determining step for denaturation of the TR as well as other retinal proteins. Unusually high thermal stability of TR multilayers, in which PSB hydrolysis is restricted due to lack of bulk water, strongly supports this proposal. Our results also show that the protonation state of the PSB counter-ion does not affect the thermal stability of the studied proteins. Thermal photo-bleaching of an artificial TR pigment derived from non-isomerizable trans-locked retinal suggests, rather counterintuitively, that the photoinduced retinal trans-cis isomerization is not a pre-requisite for light catalyzed thermal denaturation of TR. Protein conformation alteration triggered by light-induced retinal excited state formation is likely to facilitate the PSB hydrolysis.

Tuning of hyperpolarizability, and one- and two-photon absorption of donor-acceptor and donor-acceptor-acceptor-type intramolecular charge transfer-based sensors.

The present work aims to study the effect of solvent as well as arrangement of donor-acceptor groups on linear and non-linear optical (NLO) response properties of two experimentally studied intramolecular charge-transfer (ICT)-based fluorescent sensors. One of them (molecule 1) is a donor-acceptor (D-A) system with hemicyanine and dimethylanilino as electron withdrawing and donating groups, respectively, while the other one (molecule 3) is molecule 1 fused with a boron-dipyrromethene (BODIPY) moiety. BODIPY acts as the electron acceptor group of molecule 2 that as well consists of dimethylanilino as the electron donor. Density functional theory (DFT) as well as time-dependent DFT has been employed to optimize the geometry of the molecules, followed by computation of dipole moment (µ), static first hyperpolarizability (ßtotal), and one- and two-photon absorption (TPA) strengths. The results reveal that dipole moment as well as total static first hyperpolarizability (ßtotal) of the studied molecules is dominated by the respective components in the direction of charge transfer. The ratio of vector component of first hyperpolarizability (ßvec) to ßtotal also supports the unidirectional charge transfer in the studied systems. In molecule 3, which is a donor-acceptor-acceptor (D-A-A)-type system, the BODIPY moiety is found to play a major role in controlling the NLO response over the other acceptor group. Solvents are also found to play an important role in controlling the linear as well as NLO response of the studied systems. A significant increase in the first hyperpolarizability as well as TPA cross-section of the studied molecules is predicted due to an increase in the dielectric constant of the medium. The results presented are expected to provide a clue in tuning the NLO response of many ICT-based chromophores, especially those with D-A-A arrangements.

Retinal-Salinixanthin Interactions in a Thermophilic Rhodopsin.

In microbial rhodopsins (also called retinal proteins), the retinal chromophore is used for harvesting light. A carotenoid molecule has been reported to complement the retinal as light harvesting antenna in bacterial retinal proteins, although examples are scarce. In this paper, we present the formation of a novel antenna complex between thermophilic rhodopsin (TR) and the carotenoid salinixanthin (Sal). The complex formation and its structure were studied using UV-visible absorption as well as circular dichroism (CD) spectroscopies. Our studies indicate that the complex is formed in both the trimeric and monomeric forms of TR. CD spectroscopy suggests that excitonic coupling takes place between retinal and Sal. The binding of Sal with artificial TR pigments derived from synthetic retinal analogues further supports the contribution of the retinal chromophore to the CD spectrum. These studies further support the possibility of interaction between the 4-keto ring of the Sal and the retinal in TR-Sal complexes. Temperature-dependent CD spectra indicate that the positive band (ca. 482 nm) of the bisignate CD spectra of the studied complexes originates from the contribution of excitonic coupling and induced chirality of Sal in the protein binding site. The presence of a relatively smaller glycine residue in the vicinity of the retinal chromophore in TR is proposed to be crucial for binding with Sal. The results are expected to shed light on the mechanism of retinal-carotenoid interactions in other biological systems.

Interplay of twist angle and solvents with two-photon optical channel interference in aryl-substituted BODIPY dyes.

Channel interference plays a crucial role in understanding the physics behind multiphoton absorption processes. In this work, we study the role of channel interference and solvent effects on the two-photon absorption in aryl-substituted boron dipyrromethene (BODIPY) dyes, a class of intramolecular charge-transfer (ICT) molecules. For this purpose, we consider fourteen dyes of this class with various donor/acceptor substitutions at the para position of the phenyl ring and with or without methyl (-CH3) substitution on the BODIPY moiety. The presence of a methyl group on the BODIPY moiety affects the dihedral angle significantly, which in turn affects the one- (OPA) and two-photon absorption (TPA) properties of the molecules. Among the molecules studied, the one having the strong electron-donating dimethylamino group and no methyl substitution at the BODIPY moiety is found to have the highest TPA cross section. Our few-state model analysis shows that the large TPA activity of this molecule is due to the all positive contributions from different channel interference terms. Change in dielectric constant of the medium is found to have a profound impact on both the magnitude and sign of the channel interference terms. The magnitude of destructive channel interference gradually decreases with decreasing solvent polarity and becomes constructive in a low-polarity solvent. We also study the effect of rotating the phenyl ring with respect to the BODIPY moiety on the TPA activity. In the gas phase and in different solvents, we found that channel interference is changed from destructive to constructive on twisting the molecule. These results are explained by considering different dipole-, energy- and angle-terms appearing in the expression of a two-state model.

Temperature Independence of Ultrafast Photoisomerization in Thermophilic Rhodopsin: Assessment versus Other Microbial Proton Pumps.

Primary photochemical events in the unusually thermostable proton pumping rhodopsin of Thermus thermophilus bacterium (TR) are reported for the first time. Internal conversion in this protein is shown to be significantly faster than in bacteriorhodopsin (BR), making it the most rapidly isomerizing microbial proton pump known. Internal conversion (IC) dynamics of TR and BR were recorded from room temperature to the verge of thermal denaturation at 70 °C and found to be totally independent of temperature in this range. This included the well documented multiexponential nature of IC in BR, suggesting that assignment of this to ground state structural inhomogeneity needs revision. TR photodynamics were also compared with that of the phylogenetically more similar proton pump Gloeobacter rhodopsin (GR). Despite this similarity GR has poor thermal stability, and the excited state decays significantly more slowly and exhibits very prominent stretched exponential behavior. Coherent torsional wave-packets induced by impulsive photoexcitation of TR and GR show marked resemblance to each other in frequency and amplitude and differ strikingly from similar signatures in pump-probe data of BR and other microbial retinal proteins. Possible correlations between IC rates and thermal stability and the promise of using torsional coherence signatures for understanding chromophore protein binding in microbial retinal proteins are discussed.

Spectral response of 4-methyl-2,6-dicarbomethoxyphenol, an excited-state intramolecular proton-transfer probe in cyclohexane-ethanol mixtures: signatures of medium microheterogeneity.

In this paper, we explore the role of microscopic heterogeneity of the medium on the spectral response of an excited-state proton-transfer (ESIPT) probe, namely, 4-methyl-2,6-dicarbomethoxyphenol (CMOH) using steady-state and time-resolved emission spectroscopy. The mixtures of two solvents with widely different properties, viz., cyclohexane, a nonpolar, and ethanol, a polar protic solvent, were used as microheterogeneous media for spectroscopic studies. Dual fluorescence (normal and tautomer fluorescence) is observed in the nonpolar solvent (cyclohexane), while only a single peak is observed in the protic solvent, ethanol. The spectral responses of CMOH in the binary mixtures have been found to be dependent on the solvent composition and excitation wavelength. The emission spectral properties of CMOH in the cyclohexane-ethanol mixture have been seen to be superposition of spectral properties in their bulk counterparts, indicating the presence of microscopic heterogeneity in the system. A zwitterionic species of CMOH appears to have been detected in binary solvent mixtures with higher ethanol content only through low-energy excitations. The species is converted into an anionic species as excitation energy increases. Density functional theory calculations indicate that two intramolecularly hydrogen bonded rotamers of CMOH have a small energy difference. The formation of a hydrogen bonded 1:1 molecular cluster of CMOH with ethanol has been investigated in the ground state at the same level of theory. Our findings are expected to shed light on the mechanism of many acid-base reactions occurring in microscopically inhomogeneous media that often mimic many biologically relevant processes.

Spectral signatures of intramolecular charge transfer process in beta-enaminones: a combined experimental and theoretical analysis.

In this paper, we present spectroscopic signatures of intramolecular charge transfer (ICT) and effects of solvent on the ICT process in 3-(phenylamino)-2-cyclohexen-1-one (PACO), a member of the well-known molecular family, the beta-enaminones. The dual fluorescence in the steady state emission spectra of the molecule in polar solvents indicates the occurrence of ICT, which is further supported by time-resolved studies, using time correlated single photon counting technique with picosecond resolution. To understand the nature of the charge transfer, pH dependent studies of the probe in water were performed, where a quenching of fluorescence was observed even in the presence of very low concentrations of acids. Solvent induced fluorescence quenching was observed in ethanol and methanol. The ICT process was also investigated by quantum chemical calculations. To understand the role of solvents in the ICT process, we have theoretically studied the macroscopic and microscopic solvation of the probe in water. The absorption spectra of the molecule in the gas phase as well as in water were simulated using time dependent density functional theory with cc-pVTZ basis set and self-consistent reaction field theory that models macroscopic solvation. The possibility of microscopic solvation in water was probed theoretically and the formation of 1:3 molecular clusters by PACO with water molecules has been confirmed. Our findings could have a bearing on pH sensing applications of the probe.

Nanopools governing proton transfer in diametrical ways in the ground and excited state.

We present here the effects of geometrically constrained environments on the proton transfer reaction of 4-methyl 2,6-diformyl phenol (MFOH) both in the ground and excited states by employing steady-state and time-resolved fluorescence spectroscopy having picosecond and femtosecond resolutions. The nanometer-sized water pools formed in the ternary microemulsion of n-heptane-aerosol OT-water promote reprotonation of the probe. As we go on increasing the water content up to a certain value in the ground state whereas deprotonation is favored in the excited state. The emission intensity has a complex behavior as the water content is changed in the system. The lower fluidity of confined water within the reverse micelle with respect to the normal bulk water alters the related dynamics of the H-bonded network. These observations are rationalized on the basis of altered ionic water activity in the confined surroundings, i.e., on dielectric constant, ionic mobility, pH, and the favorable orientation of dipoles in the medium. Our observations might be helpful to infer about the characteristics of nanoreactors, which often mimic many biological hydrophilic pockets.