Photophysics, Molecular Reorientation in Solution and X-Ray Structure of a New Fluorescent Probe 1,7-Diazaperylene

A new fluorescent molecule 1,7-diazaperylene (DP) has been investigated by means of time-resolved and steady-state polarized fluorescence spectroscopy, as well as X-ray spectroscopy. Absorption and fluorescence spectra of DP in solution are similar to those of perylene. However, absorption and fluorescence spectra of 2,8-dimethoxy DP and 2,8-dipentyloxy DP in solution are red-shifted by ca. 55 nm relative to perylene. The fluorescence decay of DP is exponential with a lifetime of 5.1 ns in ethanol, 4.9 ns in glycerol and 4.3 ns in paraffin oil. The radiative lifetime in ethanol was calculated to be 6.3 ns for DP, 8.0 ns for 2,8-dimethoxy DP and 7.6 ns for 2,8-dipentyloxy DP. The calculated fluorescence quantum yields of 0.8 for DP and its alkoxy derivatives in ethanol, are in good agreement with those obtained from measurements. The calculated Förster radius is 37.2 ± 1 Å for DP and 41.9 ± 1 Å for its alkoxy derivatives in ethanol. Examining the S0 S1 transition, we obtain a limiting fluorescence anisotropy of r0 0.38 for DP and its alkoxy derivatives. The rotational rates of DP in paraffin oil and glycerol were compared to that of perylene. In paraffin oil both molecules show an almost identical biexponential decay of the fluorescence anisotropy, which is compatible with a rotational motion like an oblate ellipsoid. The fluorescence anisotropy is monoexponential for DP in glycerol, and DP appears to rotate like a spherical particle while perylene in glycerol appears to rotate like an oblate ellipsoid. Moreover, the rotational diffusion constant, corresponding to rotation about an axis in the aromatic plane (D), is the same for both DP and perylene in glycerol

In silico simulation of reversible and irreversible swelling of mitochondria: The role of membrane rigidity

Mitochondria have been widely accepted as the main source of ATP in the cell. The inner mitochondrial membrane (IMM) is important for the maintenance of ATP production and other functions of mitochondria. The electron transport chain (ETC) generates an electrochemical gradient of protons known as the proton-motive force across the IMM and thus produces the mitochondrial membrane potential that is critical to ATP synthesis. One of the main factors regulating the structural and functional integrity of the IMM is the changes in the matrix volume. Mild (reversible) swelling regulates mitochondrial metabolism and function; however, excessive (irreversible) swelling causes mitochondrial dysfunction and cell death. The central mechanism of mitochondrial swelling includes the opening of non-selective channels known as permeability transition pores (PTPs) in the IMM by high mitochondrial Ca2+ and reactive oxygen species (ROS). The mechanisms of reversible and irreversible mitochondrial swelling and transition between these two states are still unknown. The present study elucidates an upgraded biophysical model of reversible and irreversible mitochondrial swelling dynamics. The model provides a description of the PTP regulation dynamics using an additional differential equation. The rigidity tensor was used in numerical simulations of the mitochondrial parameter dynamics with different initial conditions defined by Ca2+ concentration in the sarco/endoplasmic reticulum. We were able to estimate the values of the IMM rigidity tensor components by fitting the model to the previously reported experimental data. Overall, the model provides a better description of the reversible and irreversible mitochondrial swelling dynamics.Funding Agency USA NIGMS NIH SC1GM128210 Institute for Functional Nanomaterials (USA NSF) 1002410 PR NASA EPSCoR (USA NASA Cooperative Agreement) NNX15AK43Ainfo:eu-repo/semantics/publishedVersio

High resolution spectroscopy of methyltrioxorhenium: towards the observation of parity violation in chiral molecules

Originating from the weak interaction, parity violation in chiral molecules has been considered as a possible origin of the biohomochirality. It was predicted in 1974 but has never been observed so far. Parity violation should lead to a very tiny frequency difference in the rovibrational spectra of the enantiomers of a chiral molecule. We have proposed to observe this predicted frequency difference using the two photon Ramsey fringes technique on a supersonic beam. Promising candidates for this experiment are chiral oxorhenium complexes, which present a large effect, can be synthesized in large quantity and enantiopure form, and can be seeded in a molecular beam. As a first step towards our objective, a detailed spectroscopic study of methyltrioxorhenium (MTO) has been undertaken. It is an ideal test molecule as the achiral parent molecule of chiral candidates for the parity violation experiment. For the 187Re MTO isotopologue, a combined analysis of Fourier transform microwave and infrared spectra as well as ultra-high resolution CO2 laser absorption spectra enabled the assignment of 28 rotational lines and 71 rovibrational lines, some of them with a resolved hyperfine structure. A set of spectroscopic parameters in the ground and first excited state, including hyperfine structure constants, was obtained for the antisymmetric Re=O stretching mode of this molecule. This result validates the experimental approach to be followed once a chiral derivative of MTO will be synthesized, and shows the benefit of the combination of several spectroscopic techniques in different spectral regions, with different set-ups and resolutions. First high resolution spectra of jet-cooled MTO, obtained on the set-up being developed for the observation of molecular parity violation, are shown, which constitutes a major step towards the targeted objective.Comment: 20 pages, 6 figure

Membrane fusion of secretory vesicles and liposomes Two different types of fusion

Secretory vesicles isolated from adrenal medulla were found to fuse in vitro in response to incubation with Ca2+. Intervesicular fusion was detected by electron microscopy and was indicated by the appearance of twinned vesicles in freeze-fractured suspensions of vesicles and in thin-sectioned pellet. Two types of fusion could be distinguished: Type I, occurring between 10−7 M and 10−4 M Ca2+, was specific for Ca2+, was inhibited by other divalent cations and was abolished by pretreatment of vesicles with glutaraldehyde, neuraminidase or trypsin. Fusion type I was linear with temperature. A second type of intervesicular fusion was elicited by Ca2+ in concentrations higher than 2.5 mM and was morphologically characterized by multiple fusions of secretory vesicles. This type of fusion was found to be similar to fusion of liposomes prepared from the membrane lipids of adrenal medullary secretory vesicles: Ca2+ could be replaced by other divalent cations, the effect of different divalent cations was additive and pretreatments attacking membrane proteins were ineffective. Fusion type II of intact secretory vesicles as well as liposome fusion was discontinuous with temperature. Liposome fusion could be detected within 35 ms and persisted for 180 min. Using liposomes containing defined Ca2+ concentrations we have not found a major influence of Ca2+ asymmetry on fusion. Incorporation of the ganglioside GM3, which is present in the membranes of intact adrenal medullary secretory vesicles did not change the properties of liposomes fusion. Using a Ca2+-selective electrode we have identified in secretory vesicle membranes both high affinity binding sites for Ca2+ (Kd = 1.6 · 10−6M) and low affinity sites (Kd = 1.2 · 10−4M)

Cholesterol Corrects Altered Conformation of MHC-II Protein in Leishmania donovani Infected Macrophages: Implication in Therapy

Previously we reported that Kala-azar patients show progressive decrease in serum cholesterol as a function of splenic parasite burden. Splenic macrophages (MΦ) of Leishmania donovani (LD) infected mice show decrease in membrane cholesterol, while LD infected macrophages (I-MΦ) show defective T cell stimulating ability that could be corrected by liposomal delivery of cholesterol. T helper cells recognize peptide antigen in the context of class II MHC molecule. It is known that the conformation of a large number of membrane proteins is dependent on membrane cholesterol. In this investigation we tried to understand the influence of decreased membrane cholesterol in I-MΦ on the conformation of MHC-II protein and peptide-MHC-II stability, and its bearing on the antigen specific T-cell activatio