The BR-body proteome contains a complex network of protein-protein and protein-RNA interactions.

Bacterial ribonucleoprotein bodies (BR-bodies) are non-membrane-bound structures that facilitate mRNA decay by concentrating mRNA substrates with RNase E and the associated RNA degradosome machinery. However, the full complement of proteins enriched in BR-bodies has not been defined. Here, we define the protein components of BR-bodies through enrichment of the bodies followed by mass spectrometry-based proteomic analysis. We find 111 BR-body-enriched proteins showing that BR-bodies are more complex than previously assumed. We identify five BR-body-enriched proteins that undergo RNA-dependent phase separation in vitro with a complex network of condensate mixing. We observe that some RNP condensates co-assemble with preferred directionality, suggesting that RNA may be trafficked through RNP condensates in an ordered manner to facilitate mRNA processing/decay, and that some BR-body-associated proteins have the capacity to dissolve the condensate. Altogether, these results suggest that a complex network of protein-protein and protein-RNA interactions controls BR-body phase separation and RNA processing.

Highly Photostable and Two-Photon Active Quantum Dot-Polymer Multicolor Hybrid Coacervate Droplets.

Fabrication and precise control of the physicochemical properties of multifunctional organic-inorganic hybrid nanocomposites find great importance in various research fields. Herein, we report the fabrication of a new class of luminescent hybrid coacervate droplets from CdTe quantum dots (QDs) and a poly(diallyldimethylammonium chloride) (PDADMAC) aqueous mixture. The colloidal stability of these droplets has been explored over wide ranges of composition, pH, and ionic strength. Although these hybrid droplets are quite stable in a low-ionic-strength medium (<100 mM NaCl) and neutral/basic pH (pH >6.5), they are unstable in a higher-ionic-strength medium (>100 mM NaCl) and acidic pH (pH <5.5). Our findings indicate specific electrostatic interactions between negatively charged QDs and positively charged PDADMAC behind the observed coacervation. They exhibit the preferential sequestration of organic dyes and serum albumins. The intrinsic luminescent properties of these hybrid droplets have been explored using confocal laser scanning microscopy (CLSM) and epifluorescence microscopy. CLSM reveals the formation of intrinsically luminescent hybrid droplets. In addition, mixed two-color luminescent droplets have been fabricated by simultaneously mixing green- and red-emitting QDs with PDADMAC aqueous solution. Epifluorescence imaging reveals highly photostable and nonbleaching photoluminescence (PL) from individual droplets as a consequence of efficient surface passivation by polymeric chains of PDADMAC. Moreover, using two-photon (2P) confocal imaging we have shown that these hybrid droplets are ideal candidates for 2P confocal imaging applications. The present study can be easily extended to fabricate a wide range of hybrid droplets with various inorganic counterparts having unique optoelectronic properties, which will further expand their applicability in nanocatalysis, bioimaging, and biosensing.

Surfactant-Induced Self-Assembly of CdTe Quantum Dots into Multicolor Luminescent Hybrid Vesicles.

Here, we report the interaction of mercaptosuccinic acid (MSA)-capped CdTe quantum dots (QDs) with hexadecyltrimethylammonium bromide (CTAB) surfactant and subsequent formation of self-assembled multicolor luminescent vesicles in aqueous medium. A continuous phase sequence from clear (C1) to turbid (T1), precipitate (P), turbid (T2), and clear (C2) has been observed for QD solution upon increasing the concentration of positively charged CTAB, indicating dynamic equilibrium between various self-assembled supramolecular structures. In contrast, no such changes have been observed in the presence of negatively charged sodium dodecyl sulfate and neutral Triton X-100 surfactants, indicating specific electrostatic interactions behind the observed phase separation behavior. Epi-fluorescence imaging in the C1 and C2 regions reveals the presence of surfactant-induced aggregates of QD. The morphologies and photoluminescence properties of self-assembled supramolecular structures in the T1 and T2 region have been explored by using scanning electron microscopy (SEM), atomic force microscopy (AFM), and confocal laser scanning microscopy (CLSM). SEM and AFM images reveal distinct spherical vesicles in the T1 and T2 regions of the binary mixture. Moreover, CLSM results show that these spherical vesicles are inherently luminescent due to the presence of self-assembled QDs. Fabrication of multicolor luminescent vesicles has been demonstrated by tuning the size of CdTe QD. Using CLSM, we have further demonstrated efficient encapsulation of Rhodamine 6G dye into these self-assembled vesicles without any structural disruption. While these luminescent vesicles are quite stable in neutral and basic pH (pH = 6.5-11), they are unstable in acidic pH (pH = 4.5-5.5). Moreover, it has been observed that this pH-responsive structural change is totally reversible. The present findings of self-assembled luminescent vesicles from QD-CTAB binary mixture may open up new opportunities in various applications such as bioimaging, drug delivery, and sensing.

Tuning of resonance energy transfer from 4',6-diamidino-2-phenylindole to an ultrasmall silver nanocluster across the lipid bilayer.

Herein we have developed a simple liposome-based donor-acceptor system across the lipid bilayer using 4',6-diamidino-2-phenylindole (DAPI) as the donor and an ultrasmall ligand-capped silver nanocluster (Ag NC) as the acceptor. The process of Förster resonance energy transfer (FRET) between DAPI and the Ag NC across the liposome bilayer has been demonstrated using steady-state and time-resolved fluorescence spectroscopy. The synthesized Ag NCs with a majority of Ag4 and Ag5 cores have been characterized using FTIR, mass spectrometry, HRTEM, UV-Vis and PL spectroscopy. Scanning electron microscopy (SEM) and dynamic light scattering (DLS) measurements reveal that the synthesized liposomes are small unilamellar vesicles (SUV) with a mean hydrodynamic diameter of 86.91 ± 6.41 nm. By using two distinct synthetic methods, we have been able to identify two selective binding sites of DAPI with liposome namely, the liposome surface and hydrophilic aqueous core. Fluorescence confocal microscopy confirms the location of the donor DAPI within different locations of liposome. It has been observed that the association of DAPI at the surface of liposome results in less efficient energy transfer to Ag NCs compared to that in the bulk aqueous medium. Energy transfer efficiency decreases from a value of 0.76 in bulk aqueous medium to a value of 0.39 for surface-associated DAPI. On the other hand encapsulation of DAPI into the hydrophilic aqueous core of a liposome results in complete inhibition of the FRET process as a consequence of increased separation distance beyond the FRET range. Hence, our study illustrates that the present DAPI-Ag NC pair can be used as a FRET marker to explore various fundamental processes across the cell membrane.

Interplay among Electrostatic, Dispersion, and Steric Interactions: Spectroscopy and Quantum Chemical Calculations of π-Hydrogen Bonded Complexes.

π-Hydrogen bonding interactions are ubiquitous in both materials and biology. Despite their relatively weak nature, great progress has been made in their investigation by experimental and theoretical methods, but this becomes significantly more complicated when secondary intermolecular interactions are present. In this study, the effect of successive methyl substitution on the supramolecular structure and interaction energy of indole⋅⋅⋅methylated benzene (ind⋅⋅⋅n-mb, n=1-6) complexes is probed through a combination of supersonic jet experiments and benchmark-quality quantum chemical calculations. It is demonstrated that additional secondary interactions introduce a subtle interplay among electrostatic and dispersion forces, as well as steric repulsion, which fine-tunes the overall structural motif. Resonant two-photon ionization and IR-UV double-resonance spectroscopy techniques are used to probe jet-cooled ind⋅⋅⋅n-mb (n=2, 3, 6) complexes, with redshifting of the N-H IR stretching frequency showing that increasing the degree of methyl substitution increases the strength of the primary N-H⋅⋅⋅π interaction. Ab initio harmonic frequency and binding energy calculations confirm this trend for all six complexes. Electronic spectra of the three dimers are broad and structureless, with quantum chemical calculations revealing that this is likely to be due to multiple tilted conformations of each dimer possessing similar stabilization energies.

Experimental observation of structures with subtle balance between strong hydrogen bond and weak n → π(*) interaction: Gas phase laser spectroscopy of 7-azaindole⋯fluorosubstituted pyridines.

In this study, interplay between a strong hydrogen bond and a very weak n → π(*) interaction has been probed through experiment for the first time. We have used resonant 2-photon ionization, Infrared-ultraviolet double resonance spectroscopy, and quantum chemistry calculation to determine the structures of 7-azaindole⋯2,6-difluoropyridine and 7-azaindole⋯2,3,5,6-tetrafluororpyridine complexes, which are stabilized by both hydrogen bonding and n → π(*) interaction. The structures of the complexes studied in the present work have been compared with the double hydrogen bonded (N-H⋯N and C-H⋯N) planar structure of 7-azaindole⋯2-fluoropyridine. It has been found that the strength of the N-H⋯N hydrogen bond in the 7-azaindole⋯2,6-substituted fluoropyridines is affected due to several factors. The main reason for huge reduction in the strength of this N-H⋯N hydrogen bond in these complexes is due to loss of the C-H⋯N hydrogen bond, through substitution of fluorine atoms in 2 and 6 positions, which induces major structural changes by bending the hydrogen bond and introducing the n → π(*) interaction. Effect of fluorination as well as presence of the n → π(*) interaction in these complexes also contributes to the reduction of the strength of the N-H⋯N interaction. Although it is difficult to quantify the role of the n → π(*) interaction to affect the strength of the hydrogen bond, observation of the structures, where a strong hydrogen bond and a weak n → π(*) interaction co-exist, is confirmed.