Localizing and Quantifying Carotenoids in Intact Cells and Tissues

Raman spectroscopy provides detailed information about the molecular structure of carotenoids. Advances in detector sensitivity and acquisition speed have driven the expansion of Raman spectroscopy from a bulk analytical tool to a powerful method for mapping carotenoid abundance in cells and tissues. In many applications, the technique is compatible with living organisms, providing highly specific molecular structure information in intact cells and tissues with subcellular spatial resolution. This leads to spatial-temporal-chemical resolution critical to understanding the complex processes in the life cycle of carotenoids and other biomolecules

Detection of High Energy Ionizing Radiation using Deeply Depleted Graphene-Oxide-Semiconductor Junctions

Graphene's linear bandstructure and two-dimensional density of states provide an implicit advantage for sensing charge. Here, these advantages are leveraged in a deeply depleted graphene-oxide-semiconductor (D2GOS) junction detector architecture to sense carriers created by ionizing radiation. Specifically, the room temperature response of the silicon-based D2GOS junction is analyzed during irradiation with 20 MeV Si4+ ions. Detection was demonstrated for doses ranging from 12-1200 ions with device functionality maintained with no substantive degradation. To understand the device response, D2GOS pixels were characterized post-irradiation via a combination of electrical characterization, Raman spectroscopy, and photocurrent mapping. This combined characterization methodology underscores the lack of discernible damage caused by irradiation to the graphene while highlighting the nature of interactions between the incident ions and the silicon absorber.Comment: 15 pages, 4 figure

Photo-physics and electronic structure of lateral graphene/MoS2 and metal/MoS2 junctions

Integration of semiconducting transition metal dichalcogenides (TMDs) into functional optoelectronic circuitries requires an understanding of the charge transfer across the interface between the TMD and the contacting material. Here, we use spatially resolved photocurrent microscopy to demonstrate electronic uniformity at the epitaxial graphene/molybdenum disulfide (EG/MoS2) interface. A 10x larger photocurrent is extracted at the EG/MoS2 interface when compared to metal (Ti/Au) /MoS2 interface. This is supported by semi-local density-functional theory (DFT), which predicts the Schottky barrier at the EG/MoS2 interface to be ~2x lower than Ti/MoS2. We provide a direct visualization of a 2D material Schottky barrier through combination of angle resolved photoemission spectroscopy with spatial resolution selected to be ~300 nm (nano-ARPES) and DFT calculations. A bending of ~500 meV over a length scale of ~2-3 micrometer in the valence band maximum of MoS2 is observed via nano-ARPES. We explicate a correlation between experimental demonstration and theoretical predictions of barriers at graphene/TMD interfaces. Spatially resolved photocurrent mapping allows for directly visualizing the uniformity of built-in electric fields at heterostructure interfaces, providing a guide for microscopic engineering of charge transport across heterointerfaces. This simple probe-based technique also speaks directly to the 2D synthesis community to elucidate electronic uniformity at domain boundaries alongside morphological uniformity over large areas

Stereodifferentiation in the intramolecular singlet excited state quenching of hydroxybiphenyl-tryptophan dyads

The photochemical processes occurring in diastereomeric dyads (S, S)-1 and (S, R)-1, prepared by conjugation of (S)-2-(2-hydroxy-1,1'-biphenyl-4-yl) propanoic acid ((S)-BPOH) with (S)- and (R)-Trp, have been investigated. In acetonitrile, the fluorescence spectra of (S, S)-1 and (S, R)-1 were coincident in shape and position with that of (S)-BPOH, although they revealed a markedly stereoselective quenching. Since singlet energy transfer from BPOH to Trp is forbidden (5 kcal mol(-1) uphill), the quenching was attributed to thermodynamically favoured (according to Rehm-Weller) electron transfer or exciplex formation. Upon addition of 20% water, the fluorescence quantum yield of (S)-BPOH decreased, while only minor changes were observed for the dyads. This can be explained by an enhancement of the excited state acidity of (S)-BPOH, associated with bridging of the carboxy and hydroxy groups by water, in agreement with the presence of water molecules in the X-ray structure of (S)-BPOH. When the carboxy group was not available for coordination with water, as in the methyl ester (S)-BPOHMe or in the dyads, this effect was prevented; accordingly, the fluorescence quantum yields did not depend on the presence or absence of water. The fluorescence lifetimes in dry acetonitrile were 1.67, 0.95 and 0.46 ns for (S)-BPOH, (S, S)-1 and (S, R)-1, respectively, indicating that the observed quenching is indeed dynamic. In line with the steady-state and time-resolved observations, molecular modelling pointed to a more favourable geometric arrangement of the two interacting chromophores in (S, R)-1. Interestingly, this dyad exhibited a folded conformation in the solid state.Financial support from the Spanish Government (CTQ2010-14882, BES-2008-003314, JCI-2011-09926, PR2011-0581), from the Generalitat Valenciana (Prometeo 2008/090) and from the Universitat Politecnica de Valencia (PAID 05-11, 2766) is gratefully acknowledged.Bonancía Roca, P.; Vayá Pérez, I.; Markovitsi, D.; Gustavsson, T.; Jiménez Molero, MC.; Miranda Alonso, MÁ. (2013). Stereodifferentiation in the intramolecular singlet excited state quenching of hydroxybiphenyl-tryptophan dyads. Organic and Biomolecular Chemistry. 11(12):1958-1963. https://doi.org/10.1039/c3ob27278hS195819631112Jiménez, M. C., Pischel, U., & Miranda, M. A. (2007). Photoinduced processes in naproxen-based chiral dyads. Journal of Photochemistry and Photobiology C: Photochemistry Reviews, 8(3), 128-142. doi:10.1016/j.jphotochemrev.2007.10.001Abad, S., Pischel, U., & Miranda, M. A. (2005). Wavelength-Dependent Stereodifferentiation in the Fluorescence Quenching of Asymmetric Naphthalene-Based Dyads by Amines. The Journal of Physical Chemistry A, 109(12), 2711-2717. doi:10.1021/jp047996aAbad, S., Vayá, I., Jiménez, M. C., Pischel, U., & Miranda, M. A. (2006). Diastereodifferentiation of Novel Naphthalene Dyads by Fluorescence Quenching and Excimer Formation. ChemPhysChem, 7(10), 2175-2183. doi:10.1002/cphc.200600337Bonancía, P., Vayá, I., Climent, M. J., Gustavsson, T., Markovitsi, D., Jiménez, M. C., & Miranda, M. A. (2012). Excited-State Interactions in Diastereomeric Flurbiprofen–Thymine Dyads. The Journal of Physical Chemistry A, 116(35), 8807-8814. doi:10.1021/jp3063838Paris, C., Encinas, S., Belmadoui, N., Climent, M. J., & Miranda, M. A. (2008). Photogeneration of 2-Deoxyribonolactone in Benzophenone−Purine Dyads. Formation of Ketyl−C1′ Biradicals. Organic Letters, 10(20), 4409-4412. doi:10.1021/ol801514vBelmadoui, N., Encinas, S., Climent, M. J., Gil, S., & Miranda, M. A. (2006). Intramolecular Interactions in the Triplet Excited States of Benzophenone–Thymine Dyads. Chemistry - A European Journal, 12(2), 553-561. doi:10.1002/chem.200500345Lhiaubet-Vallet, V., Boscá, F., & Miranda, M. A. (2007). Stereodifferentiating Drug−Biomolecule Interactions in the Triplet Excited State:  Studies on Supramolecular Carprofen/Protein Systems and on Carprofen−Tryptophan Model Dyads. The Journal of Physical Chemistry B, 111(2), 423-431. doi:10.1021/jp066968kVayá, I., Pérez-Ruiz, R., Lhiaubet-Vallet, V., Jiménez, M. C., & Miranda, M. A. (2010). Drug–protein interactions assessed by fluorescence measurements in the real complexes and in model dyads. Chemical Physics Letters, 486(4-6), 147-153. doi:10.1016/j.cplett.2009.12.091Seedher, N., & Bhatia, S. (2005). Mechanism of interaction of the non-steroidal antiinflammatory drugs meloxicam and nimesulide with serum albumin. Journal of Pharmaceutical and Biomedical Analysis, 39(1-2), 257-262. doi:10.1016/j.jpba.2005.02.031SEEDHER, N., & BHATIA, S. (2006). Reversible binding of celecoxib and valdecoxib with human serum albumin using fluorescence spectroscopic technique. Pharmacological Research, 54(2), 77-84. doi:10.1016/j.phrs.2006.02.008Nanda, R. K., Sarkar, N., & Banerjee, R. (2007). Probing the interaction of ellagic acid with human serum albumin: A fluorescence spectroscopic study. Journal of Photochemistry and Photobiology A: Chemistry, 192(2-3), 152-158. doi:10.1016/j.jphotochem.2007.05.018Zhou, B., Li, R., Zhang, Y., & Liu, Y. (2008). Kinetic analysis of the interaction between amphotericin B and human serum albumin using surface plasmon resonance and fluorescence spectroscopy. Photochemical & Photobiological Sciences, 7(4), 453. doi:10.1039/b717897bVahedian-Movahed, H., Saberi, M. R., & Chamani, J. (2011). Comparison of Binding Interactions of Lomefloxacin to Serum Albumin and Serum Transferrin by Resonance Light Scattering and Fluorescence Quenching Methods. Journal of Biomolecular Structure and Dynamics, 28(4), 483-502. doi:10.1080/07391102.2011.10508590Katrahalli, U., Kalalbandi, V. K. A., & Jaldappagari, S. (2012). The effect of anti-tubercular drug, ethionamide on the secondary structure of serum albumins: A biophysical study. Journal of Pharmaceutical and Biomedical Analysis, 59, 102-108. doi:10.1016/j.jpba.2011.09.013El-Kemary, M., Gil, M., & Douhal, A. (2007). Relaxation Dynamics of Piroxicam Structures within Human Serum Albumin Protein. Journal of Medicinal Chemistry, 50(12), 2896-2902. doi:10.1021/jm061421fTormo, L., Organero, J. A., Cohen, B., Martin, C., Santos, L., & Douhal, A. (2008). Dynamical and Structural Changes of an Anesthetic Analogue in Chemical and Biological Nanocavities. The Journal of Physical Chemistry B, 112(43), 13641-13647. doi:10.1021/jp803083yTardioli, S., Lammers, I., Hooijschuur, J.-H., Ariese, F., van der Zwan, G., & Gooijer, C. (2012). Complementary Fluorescence and Phosphorescence Study of the Interaction of Brompheniramine with Human Serum Albumin. The Journal of Physical Chemistry B, 116(24), 7033-7039. doi:10.1021/jp300055cVayá, I., Jiménez, M. C., & Miranda, M. A. (2007). Excited-State Interactions in Flurbiprofen−Tryptophan Dyads. The Journal of Physical Chemistry B, 111(31), 9363-9371. doi:10.1021/jp071301zCallis, P. R., & Burgess, B. K. (1997). Tryptophan Fluorescence Shifts in Proteins from Hybrid Simulations:  An Electrostatic Approach. The Journal of Physical Chemistry B, 101(46), 9429-9432. doi:10.1021/jp972436fLakowicz, J. R. (2000). On Spectral Relaxation in Proteins†¶‖. Photochemistry and Photobiology, 72(4), 421. doi:10.1562/0031-8655(2000)0722.0.co;2Schuler, B., & Eaton, W. A. (2008). Protein folding studied by single-molecule FRET. Current Opinion in Structural Biology, 18(1), 16-26. doi:10.1016/j.sbi.2007.12.003Shen, X., & Knutson, J. R. (2001). Subpicosecond Fluorescence Spectra of Tryptophan in Water. The Journal of Physical Chemistry B, 105(26), 6260-6265. doi:10.1021/jp010384vBeechem, J. M., & Brand, L. (1985). Time-Resolved Fluorescence of Proteins. Annual Review of Biochemistry, 54(1), 43-71. doi:10.1146/annurev.bi.54.070185.000355Callis, P. R. (1997). [7] 1La and 1Lb transitions of tryptophan: Applications of theory and experimental observations to fluorescence of proteins. Flourescence Spectroscopy, 113-150. doi:10.1016/s0076-6879(97)78009-1Basarić, N., & Wan, P. (2006). Competing Excited State Intramolecular Proton Transfer Pathways from Phenol to Anthracene Moieties. The Journal of Organic Chemistry, 71(7), 2677-2686. doi:10.1021/jo0524728Lukeman, M., & Wan, P. (2003). Excited-State Intramolecular Proton Transfer ino-Hydroxybiaryls:  A New Route to Dihydroaromatic Compounds. Journal of the American Chemical Society, 125(5), 1164-1165. doi:10.1021/ja029376yKeck, J., Kramer, H. E. A., Port, H., Hirsch, T., Fischer, P., & Rytz, G. (1996). Investigations on Polymeric and Monomeric Intramolecularly Hydrogen-Bridged UV Absorbers of the Benzotriazole and Triazine Class. The Journal of Physical Chemistry, 100(34), 14468-14475. doi:10.1021/jp961081hVollmer, F., & Rettig, W. (1996). Fluorescence loss mechanism due to large-amplitude motions in derivatives of 2,2′-bipyridyl exhibiting excited-state intramolecular proton transfer and perspectives of luminescence solar concentrators. Journal of Photochemistry and Photobiology A: Chemistry, 95(2), 143-155. doi:10.1016/1010-6030(95)04252-0Lukeman, M., & Wan, P. (2002). A New Type of Excited-State Intramolecular Proton Transfer:  Proton Transfer from Phenol OH to a Carbon Atom of an Aromatic Ring Observed for 2-Phenylphenol1. Journal of the American Chemical Society, 124(32), 9458-9464. doi:10.1021/ja0267831Jiménez, M. C., Miranda, M. A., Tormos, R., & Vayá, I. (2004). Characterisation of the lowest singlet and triplet excited states of S-flurbiprofen. Photochem. Photobiol. Sci., 3(11-12), 1038-1041. doi:10.1039/b408530bWeller, A. (1982). Photoinduced Electron Transfer in Solution: Exciplex and Radical Ion Pair Formation Free Enthalpies and their Solvent Dependence. Zeitschrift für Physikalische Chemie, 133(1), 93-98. doi:10.1524/zpch.1982.133.1.093Winget, P., Cramer, C. J., & Truhlar, D. G. (2004). Computation of equilibrium oxidation and reduction potentials for reversible and dissociative electron-transfer reactions in solution. Theoretical Chemistry Accounts, 112(4). doi:10.1007/s00214-004-0577-0ÇAKIR, S., & BÇER, E. (2010). SYNTHESIS, SPECTRAL CHARACTERIZATION AND ELECTROCHEMISTRY OF VANADIUM(V) COMPLEX WITH TRYPTOPHAN. Journal of the Chilean Chemical Society, 55(2). doi:10.4067/s0717-9707201000020002

Low-fidelity DNA synthesis by the L979F mutator derivative of Saccharomyces cerevisiae DNA polymerase ζ

To probe Pol ζ functions in vivo via its error signature, here we report the properties of Saccharomyces cerevisiae Pol ζ in which phenyalanine was substituted for the conserved Leu-979 in the catalytic (Rev3) subunit. We show that purified L979F Pol ζ is 30% as active as wild-type Pol ζ when replicating undamaged DNA. L979F Pol ζ shares with wild-type Pol ζ the ability to perform moderately processive DNA synthesis. When copying undamaged DNA, L979F Pol ζ is error-prone compared to wild-type Pol ζ, providing a biochemical rationale for the observed mutator phenotype of rev3-L979F yeast strains. Errors generated by L979F Pol ζ in vitro include single-base insertions, deletions and substitutions, with the highest error rates involving stable misincorporation of dAMP and dGMP. L979F Pol ζ also generates multiple errors in close proximity to each other. The frequency of these events far exceeds that expected for independent single changes, indicating that the first error increases the probability of additional errors within 10 nucleotides. Thus L979F Pol ζ, and perhaps wild-type Pol ζ, which also generates clustered mutations at a lower but significant rate, performs short patches of processive, error-prone DNA synthesis. This may explain the origin of some multiple clustered mutations observed in vivo