Probing Behavior Research Articles

Contrasting complexation behaviour of zwitterionic amyloid probe, SYPRO orange with β-cyclodextrin and captisol

The differential complexation studies with cyclodextrins and their charged derivatives have been well studied using cationic, anionic, as well as neutral guest molecules. However, supramolecular complexation with zwitterionic guest molecules is quite limited. Zwitterions are overall neutral molecules with localized charges that can offer additional site-specific electrostatic interaction with cyclodextrins as compared to neutral guest molecules. There is no information on how the different intra- and intermolecular interactions can modulate the binding of a zwitterion with a macrocyclic host. Herein, the supramolecular interactions of biologically important zwitterionic amyloid probe, SYPRO Orange (SO) with β-cyclodextrin (β-CD) and sulfobutylether β-cyclodextrin (SBE-β-CD, Captisol®) have been evaluated and compared using different spectroscopic methods. SO forms inclusion complexes with both β-CD and SBE-β-CD which led to the enhanced fluorescence along with a hypsochromic shift in its emission spectra. It has been shown that the presence of segmental rotation in a guest molecule with charge transfer property favours its association with the macromolecules. The fluorescence enhancement and the binding affinities show contrasting behavior in these two types of host–guest complexes. Time-resolved emission measurements could disentangle the contribution from different inclusion complexes. Ionic strength and temperature variation in the photophysical properties helped to understand the molecular interactions involved in such contrasting behavior.

Insight into the Sensing Behavior of DNA Probes Based on MOF-Nucleic Acid Interaction for Bioanalysis.

Adsorption of DNA probes onto nanomaterials is a promising strategy for bioassay establishment typically using fluorescence or catalytic activities to generate signals. Albeit important, there is currently a lack of systematic understanding of the sensing behaviors building on nanomaterial-DNA interactions, which greatly limits the rational method design and their subsequent applications. Herein, the issue was investigated by employing multifunctional metal-organic frameworks (MOFs) (FeTCPP⊂UiO-66) as a model that was synthesized via integrating heme-like ligand FeTCPP into commonly used MOFs (UiO-66). Our results demonstrated that the fluorescently labeled DNA adsorbed onto FeTCPP⊂UiO-66 was quenched through photoinduced electron transfer, fluorescence resonance energy transfer, and the internal filtration effect. Among different DNA structures, double-stranded DNA and hybridization chain reaction products largely retained their fluorescence due to desorption and conformational variation, respectively. In addition, ssDNA could maximally inhibit the peroxidase activity of FeTCPP⊂UiO-66, and this inhibition was strongly dependent on the strand length but independent of base composition. On the basis of these discoveries, a fluorescence/colorimetric dual-modal detection was designed against aflatoxin B1 with satisfactory performances obtained to further verify our results. This study provided some new insights into the sensing behaviors based on MOF-DNA interactions, indicating promising applications for rational bioassay design and its performance improvement.

Investigation on non-radioactive behavior of an acylhydrazone-based fluorescent probe: Coexistence of PET and TICT mechanisms

A fluorescent probe (E)-((2,4-dihydroxybenzyl)diazenyl)(pyridin-2-yl)methanone (HL) to effectively and selectively detect Al3+ was designed and synthesized in the experiment. Herein, we explained the excited state dynamics mechanism of HL by using density functional theory (DFT) and time-dependent density functional theory (TD-DFT). The potential energy surfaces (PESs) proved that the excited-state intramolecular proton transfer (ESIPT) process hardly occurs due to the high reaction barriers, so the fluorescence quenching behavior of HL was not based on ESIPT. The frontier molecular orbitals (FMOs) and spectral properties were analyzed to better understand the origination of fluorescence quenching. It was found that an electron on C = N in HL could be transferred to the fluorophore during excitation in the absence of Al3+, accompanied by the PET process. The excited state could undergo a twisted intramolecular charge transfer (TICT) process, releasing non-radiative decay. After binding to Al3+, the photo-induced electron transfer (PET) process has no longer occurred, and the TICT process is eliminated, resulting in a significant fluorescence enhancement. Therefore, the calculation results well explain the quenching and enhancement behaviors of fluorescence before and after the reaction with Al3+.

Highly pH-Responsive Sensor Based on a EuIII Metal-Organic Framework with Efficient Recognition of Arginine and Lysine in Living Cells.

A lanthanide-based three-dimensional metal-organic framework with excellent water, acid/base, and solvent stability, namely {[(CH3)2NH2]0.7[Eu2(BTDBA)1.5(lac)0.7(H2O)2]·2H2O·2DMF·2CH3CN}n (JXUST-29, H4BTDBA = 4',4‴-(benzo[c][1,2,5]thiadiazole-4,7-diyl)bis([1,1'-biphenyl]-3,5-dicarboxylic acid), Hlac = lactic acid), has been synthesized and characterized. Since the N atoms of the thiadiazole group will not coordinate with lanthanide ions, JXUST-29 has a free basic N-site accessible to small H+ ions, which allows it to be used as a promising pH fluorescence sensor. Interestingly, the luminescence signal was significantly enhanced, with an approximately 54-fold enhancement in the emission intensity when the pH value was increased from 2 to 5, which is the typical behavior of pH probes. In addition, JXUST-29 can also be used as a luminescence sensor to detect l-arginine (Arg) and l-lysine (Lys) in an aqueous solution through fluorescence enhancement and the blue-shift effect. The detection limits were 0.023 and 0.077 μM, respectively. In addition, JXUST-29-based devices were designed and developed to facilitate detection. Importantly, JXUST-29 is also capable of detecting and sensing Arg and Lys in living cells.

Strategies for Competitive Activity‐Based Protein Profiling in Small Molecule Inhibitor Discovery and Characterization

AbstractSince its introduction by Cravatt and colleagues in 1999, activity‐based protein profiling (ABPP) has become widely utilized throughout academia, government, and industry laboratories to study enzymes spanning numerous gene families in a multitude of biological systems. As a variation of ABPP, competitive ABPP provides a powerful approach to characterize the binding behavior of small molecule probes and clinical drugs throughout the functional proteome. The power and flexibility of competitive ABPP are exemplified by a wide range of creative adaptions which increase assay throughput, enable diverse detection schemes, and support the implementation of this approach within a hybrid target‐based screening platform. We review major developments in competitive ABPP through a compare‐contrast format to provide a useful introduction to this enabling technology for scientists in chemical biology and drug discovery.

Type-IV glandular trichomes disrupt the probing behavior of Bemisia tabaci MEAM1 and Tomato severe rugose virus inoculation in tomato plants

Type-IV glandular trichomes disrupt the probing behavior of Bemisia tabaci MEAM1 and Tomato severe rugose virus inoculation in tomato plants

Self-Quenching Behavior of a Fluorescent Probe Incorporated within Lipid Membranes Explored Using Electrophoresis and Fluorescence Lifetime Imaging Microscopy.

Fluorescent probes are useful in biophysics research to assess the spatial distribution, mobility, and interactions of biomolecules. However, fluorophores can undergo "self-quenching" of their fluorescence intensity at high concentrations. A greater understanding of concentration-quenching effects is important for avoiding artifacts in fluorescence images and relevant to energy transfer processes in photosynthesis. Here, we show that an electrophoresis technique can be used to control the migration of charged fluorophores associated with supported lipid bilayers (SLBs) and that quenching effects can be quantified with fluorescence lifetime imaging microscopy (FLIM). Confined SLBs containing controlled quantities of lipid-linked Texas Red (TR) fluorophores were generated within 100 × 100 μm corral regions on glass substrates. Application of an electric field in-plane with the lipid bilayer induced the migration of negatively charged TR-lipid molecules toward the positive electrode and created a lateral concentration gradient across each corral. The self-quenching of TR was directly observed in FLIM images as a correlation of high concentrations of fluorophores to reductions in their fluorescence lifetime. By varying the initial concentration of TR fluorophores incorporated into the SLBs from 0.3% to 0.8% (mol/mol), the maximum concentration of fluorophores reached during electrophoresis could be modulated from 2% up to 7% (mol/mol), leading to the reduction of fluorescence lifetime down to 30% and quenching of the fluorescence intensity down to 10% of their original levels. As part of this work, we demonstrated a method for converting fluorescence intensity profiles into molecular concentration profiles by correcting for quenching effects. The calculated concentration profiles have a good fit to an exponential growth function, suggesting that TR-lipids can diffuse freely even at high concentrations. Overall, these findings prove that electrophoresis is effective at producing microscale concentration gradients of a molecule-of-interest and that FLIM is an excellent approach to interrogate dynamic changes to molecular interactions via their photophysical state.

The cauliflower mosaic virus transmission helper protein P2 modifies directly the probing behavior of the aphid vector Myzus persicae to facilitate transmission.

There is growing evidence that plant viruses manipulate their hosts and vectors in ways that increase transmission. However, to date only few viral components underlying these phenomena have been identified. Here we show that cauliflower mosaic virus (CaMV) protein P2 modifies the feeding behavior of its aphid vector. P2 is necessary for CaMV transmission because it mediates binding of virus particles to the aphid mouthparts. We compared aphid feeding behavior on plants infected with the wild-type CaMV strain Cabb B-JI or with a deletion mutant strain, Cabb B-JIΔP2, which does not produce P2. Only aphids probing Cabb B-JI infected plants doubled the number of test punctures during the first contact with the plant, indicating a role of P2. Membrane feeding assays with purified P2 and virus particles confirmed that these viral products alone are sufficient to cause the changes in aphid probing. The behavior modifications were not observed on plants infected with a CaMV mutant expressing P2Rev5, unable to bind to the mouthparts. These results are in favor of a virus manipulation, where attachment of P2 to a specific region in the aphid stylets-the acrostyle-exercises a direct effect on vector behavior at a crucial moment, the first vector contact with the infected plant, which is essential for virus acquisition.

The cauliflower mosaic virus transmission helper protein P2 modifies directly the probing behavior of the aphid vector Myzus persicae to facilitate transmission

The cauliflower mosaic virus transmission helper protein P2 modifies directly the probing behavior of the aphid vector Myzus persicae to facilitate transmission

Membranes with large phospholipid asymmetries.

Plasma membranes in cells are asymmetric, an observation that dates 40 years. Recent observations suggest that these membranes present large lipid number asymmetries, with almost twice as many phospholipids on one of the leaflet than the other. Simulations provide an excellent avenue to probe behavior of these membranes. Here, I discuss the behavior of such membranes, in particular with respect to chemical asymmetries in the membrane. The work required to establish the phospholipid number asymmetry is also discussed, a quantity that is directly related to the work done on lipids by the so-called flippases and floppases proteins responsible for asymmetry homeostasis.

Voltammetric examination of carbon and platinum nanoelectrodes under mixed diffusion – migration mass transport conditions

The majority of electroanalytical experiments are performed with supporting electrolytes to minimize the migration of electroactive ions and other unwanted effects of the low conductivity of the system. Microelectrodes and nanoelectrodes allow for voltammetric measurements in solutions of substantial resistivity, however, their application in non-supported electrolytes is vaguely studied in detail. This work demonstrates the employment of carbon and platinum nanoelectrodes in solutions of ionic and neutral ferrocene derivatives in the presence and absence of supporting electrolyte. The ferrocene derivatives of various charges served as redox probes for the studies of diffusion-migration transport to nanoelectrodes. The obtained results are confronted with the theoretical predictions derived for microelectrodes. An addition of a supporting electrolyte stabilizes the voltammetric signal. It makes the voltammetric waves at nanoelectrodes well developed irrespective of the type of the redox system examined. The results revealed that the migration of electroactive species to nanoelectrodes in the absence of a supporting electrolyte does not introduce significant changes to the voltammetric signal recorded with a supporting electrolyte as much as for bigger microelectrodes. In many cases, the complete elimination of supporting electrolyte led to the poor development of the voltammetric waves at nanoelectrodes, possibly due to the breakdown of the electroneutrality in the region adjacent to the nanoelectrode, since the electrical double layer for nanoelectrodes may occupy a significant fraction of the transport depletion layer. The uncertainty in the concentrations of ionic impurities (that determine the actual ionic strength) in the layer adjacent to the electrode is another factor that may produce unpredictable and, to some extent, random voltammetric behavior of charged redox probes at nanoelectrodes in the absence of supporting electrolyte.

Probing behavior of Aphis fabae and Myzus persicae on three species of grapevines with analysis of grapevine leaf anatomy and allelochemicals

The peach-potato aphid Myzus persicae (Sulzer) and the black bean aphid Aphis fabae Scopoli are polyphagous and cosmopolitan hemipterans, therefore they can infest grapevines in all areas of cultivation. Electrical Penetration Graph (EPG) technique was applied to monitor the probing behavior of A. fabae and M. persicae on Vitis amurensis Rupr., Vitis riparia Michaux, and Vitis vinifera L. The content of major flavonoids and stilbenoids in grapevine leaves and epidermal thickness, distance between abaxial leaf surface and phloem, and the simulated shortest pathway from epidermis to phloem that might have affected aphid probing were also analyzed. Aphid probing was limited mainly to non-vascular tissues on the three studied grapevine species. Phloem phase occurred in 32%, 14%, and 6% of A. fabae and in 76%, 39%, and 74% of M. persicae on V. amurensis, V. riparia and V. vinifera, respectively. Phloem phase consisted of only salivation into sieve elements and lasted less than 2.5 minutes on average in all aphids. The time to reach the first phloem phase on grapevines was 5.0 hours in A. fabae and 2.6–3.6 hours in M. persicae. Of the analyzed flavonoids, catechin, epicatechin, and quercetin occurred in all grapevine species, while rutin – in V. amurensis and V. riparia and isorhamnetin only in V. amurensis. Of the analyzed stilbenoids, piceid occurred in all grapevines, resveratrol in V. amurensis and V. vinifera, and ε-viniferin only in V. vinifera. Aphid behavior demonstrated that V. amurensis, V. riparia and V. vinifera are not attractive host plants to A. fabae and M. persicae. It is likely that the content of flavonoids and stilbenoids contributes to the limited susceptibility of the three grapevine species to A. fabae and M. persicae, while the observed slight differences in the anatomical structure of the leaves seem not significant in this context.

Tuning the aggregation-induced emission behavior of novel luminescence probes for DNP and Fe3+sensing by molecular stacking

The effect of molecular stacking on AIE behavior was explored by synthesizing a series of biscarbazole-type molecules with different substituents. The non-compact packed TPAK molecules have AIE behavior and can be used to detect DNP and Fe3+.

Fluorescence of pyrene and its derivatives to reveal constituent and composition dependent solvation within hydrophobic deep eutectic solvents.

Depending on their constituents and composition, deep eutectic solvents (DESs) may exhibit widely varying physicochemical properties. Based on the miscibility of water in a DES, it can broadly be classified as 'hydrophobic' or 'hydrophilic'. The polarity offered by hydrophobic DESs as compared to common organic solvents in the context of solute solubilization thus becomes of the utmost importance. A versatile fluorescence probe, pyrene (Py), its aldehyde derivative, pyrene-1-carboxaldehyde (PyCHO), and a termini end-tagged dipyrenyl polydimethylsiloxane polymer (Py-PDMS-Py) are employed to assess the solvation environment afforded by DESs composed of thymol (Thy), (-)-menthol (Men), and n-decanoic acid (DA). To decipher the effect of the constituents and composition on solute solvation, DESs composed of different pairs of constituents in different molar ratios, namely Thy : Men (1 : 1 and 1 : 2), DA : Men (1 : 1 and 1 : 2), and Thy : DA (2 : 1, 1 : 1, and 1 : 2), are investigated. Pyrene, through its band 1-to-band 3 emission intensity ratio (Py I1/I3), displays higher cybotactic region dipolarity in Thy-containing DESs due to the phenyl ring of Thy; the sensitivity of Py I1/I3 towards temperature is also higher in Thy DESs. The fluorescence lifetime of pyrene and its temperature dependence are higher in Men-containing DESs in comparison. Quenching of pyrene fluorescence by nitromethane in these DESs is dynamic in nature and the recovered bimolecular quenching rate constants (kq) reveal efficient diffusion of the fluorophore-quencher pair in these DESs as compared to other iso-viscous media. The kq adheres to the Stokes-Einstein relation, implying inherent homogeneity associated to these DESs. PyCHO emission spectra show a high energy structured band in Thy : Men DESs, whereas the band shifts bathochromically and becomes broad in DA-containing DESs. The PyCHO cybotactic region is relatively nonpolar in Thy : Men DESs as compared to Thy : DA and Men : DA DESs. The extent of intramolecular excimer formation by Py-PDMS-Py reveals these DESs to be "good" solvents for polymer solvation where DES-polymer interactions are maximized. The microviscosity (ημ) surrounding Py-PDMS-Py correlates with the bulk dynamic viscosity (ηbulk) within the investigated DESs, further corroborating the absence of microheterogeneity. Overall, the observations reveal the similarity of these hydrophobic DESs to common organic solvents in the context of solute solubilization.

Scalar correlators and normal modes in holographic neutron stars

The holographic neutron star provides a strong coupling description for a highly degenerate metallic state on a sphere. Its phase space can be split into two different sectors, with an unstable region at intermediate degeneracies. We investigate the critical nature of such region, by analyzing the asymptotic behavior of a scalar probe to compute the two-point correlator of the boundary theory. We show that in the stable region the correlator is dominated by the normal modes, whereas it displays a critical power-law behavior as we move into the unstable region.

Role of Acrostyle Cuticular Proteins in the Retention of an Aphid Salivary Effector

To avoid the activation of plant defenses and ensure sustained feeding, aphids are assumed to use their mouthparts to deliver effectors into plant cells. A recent study has shown that effectors detected near feeding sites are differentially distributed in plant tissues. However, the precise process of effector delivery into specific plant compartments is unknown. The acrostyle, a cuticular organ located at the tip of maxillary stylets that transiently binds plant viruses via its stylin proteins, may participate in this specific delivery process. Here, we demonstrate that Mp10, a saliva effector released into the plant cytoplasm during aphid probing, binds to the acrostyles of Acyrthosiphon pisum and Myzus persicae. The effector probably interacts with Stylin-03 as a lowered Mp10-binding to the acrostyle was observed upon RNAi-mediated reduction in Stylin-03 production. In addition, Stylin-03 and Stylin-01 RNAi aphids exhibited changes in their feeding behavior as evidenced by electrical penetration graph experiments showing longer aphid probing behaviors associated with watery saliva release into the cytoplasm of plant cells. Taken together, these data demonstrate that the acrostyle also has effector binding capacity and supports its role in the delivery of aphid effectors into plant cells.

Comparative study of neuropeptide signaling systems in Hemiptera.

Numerous physiological processes in insects are tightly regulated by neuropeptides and their receptors. Although they form an ancient signaling system, there is still a great deal of variety in neuropeptides and their receptors among different species within the same order. Neuropeptides and their receptors have been documented in many hemipteran insects, but the differences among them have been poorly characterized. Commercial grapevines worldwide are plagued by the bug Daktulosphaira vitifoliae (Hemiptera: Sternorrhyncha). Here, 33 neuropeptide precursors and 48 putative neuropeptide G protein-coupled receptor (GPCR) genes were identified in D. vitifoliae. Their expression profiles at the probe and feeding stages reflected potential regulatory roles in probe behavior. By comparison, we found that the Releasing Hormone-Related Peptides (GnRHs) system of Sternorrhyncha was differentiated from those of the other 2 suborders in Hemiptera. Independent secondary losses of the adipokinetic hormone/corazonin-related peptide receptor (ACP) and corazonin (CRZ) occurred during the evolution of Sternorrhyncha. Additionally, we discovered that the neuropeptide signaling systems of Sternorrhyncha were very different from those of Heteroptera and Auchenorrhyncha, which was consistent with Sternorrhyncha's phylogenetic position at the base of the order. This research provides more knowledge on neuropeptide systems and sets the groundwork for the creation of novel D. vitifoliae management strategies that specifically target these signaling pathways.

Bulk and confined acetonitrile in mesoporous silica matrices by extrinsic probing via ESR technique: Effects of pore topology, pore size and pore surface composition

The spectral behavior of the spin probe 2,2,6,6-tetramethyl-1‑piperidinyloxy (TEMPO) in acetonitrile (ACN) as arepresentative of aprotic polar medium in the bulk and in 2D-confined states of three silica matrices differing in the pore topology, pore size and pore wall composition using electron spin resonance (ESR) is presented. The changes in interaction and dynamic parameters of TEMPO due to spatial restriction of ACN and pore surface chemistry are related to phase behaviors from differential scanning calorimetry (DSC). The obtained ESR findings are compared with those for apolar and protic polar media in the same native and modified confining silica matrices.

Catalyzing systemic movement of inward rectifier potassium channel inhibitors for antifeedant activity against the cotton aphid, Aphis gossypii (Glover).

The cotton aphid, Aphis gossypii Glover (Hemiptera: Aphididae), is a destructive agricultural pest, capable of photosynthate removal and plant virus transmission. Therefore, we aimed to test the antifeedant properties of small-molecule inhibitors of inward rectifier potassium (Kir) channels expressed in insect salivary glands and develop an approach for enabling systemic movement of lipophilic Kir inhibitors. Two Kir channel inhibitors, VU041 and VU730, reduced the secretory activity of the aphid salivary glands by 3.3-fold and foliar applications of VU041 and VU730 significantly (P < 0.05) increased the time to first probe, total probe duration, and nearly eliminated phloem salivation and ingestion. Next, we aimed to facilitate systemic movement of VU041 and VU730 through evaluation of a novel natural product based solubilizer containing rubusoside that was isolated from Chinese sweet leaf (Rubus suavissimus) plants. A single lower leaf was treated with Kir inhibitor soluble liquid (KI-SL) and systemic movement throughout the plant was verified via toxicity bioassays and changes to feeding behavior through the electrical penetration graph (EPG) technique. EPG data indicate KI-SL significantly reduced ability to reach E1 (phloem salivation) and E2 (phloem ingestion) waveforms and altered plant probing behavior when compared to the untreated control. High-performance liquid chromatography (HPLC) analysis indicated the presence of VU041 and VU730 in the upper leaf tissue of these plants. Together, these data provide strong support that incorporation of rubusoside with Kir inhibitors enhanced translaminar and translocation movement through the plant tissue. These data further support hemipteran Kir channels as a target to prevent feeding and induce toxicity. Further, these studies highlight a novel delivery approach for generating plant systemic activity of lipophilic insecticides. © 2022 Society of Chemical Industry.

Decanoic Acid Based Deep Eutectic Solvents: Constituent-Controlled Physicochemical Properties and Solvatochromic Probe Behavior

Decanoic Acid Based Deep Eutectic Solvents: Constituent-Controlled Physicochemical Properties and Solvatochromic Probe Behavior