scholarly journals Enzymatic digestion of luminescent albumin-stabilized gold nanoclusters under anaerobic conditions: clues to the quenching mechanism

2021 ◽  
Author(s):  
Matylda Wacławska ◽  
Wojciech Dzwolak
Keyword(s):  
Gold Nanoclusters ◽  
Enzymatic Digestion ◽  
Proteinase K ◽  
Moderate Increase ◽  
Anaerobic Conditions ◽  
Previous Hypothesis ◽  
Potential Applications ◽  
Mechanisms Of Formation ◽  
Protease Treatment ◽  
Red Luminescence

Many of the potential applications of albumin-stabilized gold nanoclusters (AuNC) arise from the sensitivity of their luminescence to the presence of various ions and albumin-degrading proteases. However, the underlying photophysics and the mechanisms responsible for protease-induced quenching of AuNC luminescence are not fully understood. Here, we study proteinase K-induced digestion of bovine serum albumin (BSA)-AuNC conjugate under aerobic and anaerobic conditions. To this end, we adapt a Co(II)-catalyzed sulfite-based protocol enabling effective in situ deoxidization without deactivation of the enzyme. In the absence of proteinase K, the anaerobic conditions facilitate luminescence of BSA-AuNC reflected by a moderate increase in the red luminescence intensity. However, in the presence of proteinase K, we have observed a steeper decrease of emission intensity irrespective of whether the digestion was carried out under aerobic or anaerobic conditions. In both cases, the diminishing fluorescence occurred in phase with shifting of the emission maximum to longer wavelengths. These results contradict the previous hypothesis that protease-induced quenching of BSA-AuNC luminescence is a consequence of enhanced diffusion of oxygen to bare AuNC. Instead, aggregation of unprotected AuNCs and separation of nanoclusters from albumin’s side chains involved in energy transfers and luminescence-promoting electron donors may underlie the observed sensitivity of BSA-AuNC to protease treatment. Our findings are discussed in the context of mechanisms of formation and photophysics of BSA-AuNC conjugates.

scholarly journals Monitoring of Enzymatic Proteolysis Using Self-Assembled Quantum Dot-Protein Substrate Sensors

2008 ◽  
Vol 2008 ◽  
pp. 1-10 ◽  
Author(s):  
Aaron R. Clapp ◽  
Ellen R. Goldman ◽  
H. Tetsuo Uyeda ◽  
Eddie L. Chang ◽  
Jessica L. Whitley ◽  
...  
Keyword(s):  
Quantum Dot ◽  
Proteolytic Enzymes ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Enzymatic Digestion ◽  
Enzymatic Proteolysis ◽  
Potential Applications ◽  
Multiple Copies ◽  
Self Assembled

We have previously utilized hybrid semiconductor quantum dot- (QD-) peptide substrates for monitoring of enzymatic proteolysis. In this report, we expand on this sensing strategy to further monitor protein-protease interactions. We utilize QDs self-assembled with multiple copies of dye-labeled proteins as substrates for the sensing of protease activity. Detection of proteolysis is based on changes in the rate of fluorescence resonance energy transfer (FRET) between the QDs and the proximal dye-labeled proteins following protein digestion by added enzyme. Our study focused on two representative proteolytic enzymes: the cysteine protease papain and the serine protease endoproteinase K. Analysis of the enzymatic digestion allowed us to estimate minimal values for the enzymatic activities of each enzyme used. Mechanisms of enzymatic inhibition were also inferred from the FRET data collected in the presence of inhibitors. Potential applications of this technology include drug discovery assays and in vivo cellular monitoring of enzymatic activity.

scholarly journals Novel Tunable Green-Red Luminescence in Mn2+ Doped Ca9Tb(PO4)7 Phosphors Based on the Mn2+ Regulation and Energy Transfer

Coatings ◽  
2020 ◽  
Vol 10 (10) ◽  
pp. 952
Author(s):  
Bingwen Yang ◽  
Yefeng Feng ◽  
Qinghu Zhao ◽  
Miao He ◽  
Yang Lv
Keyword(s):  
Energy Transfer ◽  
Green Emission ◽  
Energy Transfer Mechanism ◽  
Red Emission ◽  
Broadband Emission ◽  
Characteristic Emission ◽  
Potential Applications ◽  
Light Excitation ◽  
Red Luminescence ◽  
Thermal Stability Of

β-Ca3(PO4)2 type phosphors Ca9Tb(PO4)7:Mn2+ were fabricated by high temperature solid state reaction. Under 377 nm light excitation, the Ca9Tb(PO4)7 host displays the green emission attributable to the characteristic emission of Tb3+ ions peaking at 488, 542, 586, and 620 nm, respectively. The red broadband emission is observed when Ca9Tb(PO4)7 is doped with Mn2+ ions. The emission is attributed to the energy transfer from Tb3+ to Mn2+ ions; this facilitates the realization of the tunable green–red emission. The energy transfer mechanism from Tb3+ to Mn2+ is defined as quadrupole–quadrupole interaction. Furthermore, the thermal stability of Ca9Tb(PO4)7:Mn2+ samples has been studied, and it can maintain half the emission intensity exceeding 424 K. This demonstrates their potential applications in white light LEDs (w-LEDs).

Spherical SiO2 @ GdPO4 : Eu3+ Core–Shell Phosphors: Sol–Gel Synthesis and Characterization

2007 ◽  
Vol 7 (2) ◽  
pp. 542-548 ◽  
Author(s):  
Cuikun Lin ◽  
Bo Zhao ◽  
Zhenling Wang ◽  
Min Yu ◽  
Huan Wang ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Low Voltage ◽  
Uv Light ◽  
Sol Gel ◽  
Emission Spectra ◽  
Core Shell ◽  
Time Resolved ◽  
Potential Applications ◽  
Ft Ir ◽  
Red Luminescence

Nanocrystalline GdPO4 : Eu3+ phosphor layers were coated on non-aggregated, monodisperse and spherical SiO2 particles by Pechini sol–gel method, resulting in the formation of core–shell structured SiO2 @ GdPO4 : Eu3+ particles. X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FT-IR), field emission scanning electron microscopy (FESEM), transmission electron microscopy (TEM), photoluminescence (PL), low-voltage cathodoluminescence (CL), time-resolved PL spectra and lifetimes were used to characterize the core–shell structured materials. Both XRD and FT-IR results indicate that GdPO4 layers have been successfully coated on the SiO2 particles, which can be further verified by the images of FESEM and TEM. Under UV light excitation, the SiO2 @ GdPO4 : Eu3+ phosphors show orange-red luminescence with Eu3+ 5D0–7F1 (593 nm) as the most prominent group. The PL excitation and emission spectra suggest that an energy transfer occurs from Gd3+ to Eu3+ in SiO2 @ GdPO4 : Eu3+ phosphors. The obtained core–shell phosphors have potential applications in FED and PDP devices.

scholarly journals High-viscosity injector-based pink-beam serial crystallography of microcrystals at a synchrotron radiation source

IUCrJ ◽  
2019 ◽  
Vol 6 (3) ◽  
pp. 412-425 ◽  
Author(s):  
Jose M. Martin-Garcia ◽  
Lan Zhu ◽  
Derek Mendez ◽  
Ming-Yue Lee ◽  
Eugene Chun ◽  
...  
Keyword(s):  
Synchrotron Radiation ◽  
Radiation Source ◽  
Structural Changes ◽  
High Viscosity ◽  
Synchrotron Radiation Source ◽  
Proteinase K ◽  
Moderate Increase ◽  
Time Resolved ◽  
X Ray ◽  
Serial Crystallography

Since the first successful serial crystallography (SX) experiment at a synchrotron radiation source, the popularity of this approach has continued to grow showing that third-generation synchrotrons can be viable alternatives to scarce X-ray free-electron laser sources. Synchrotron radiation flux may be increased ∼100 times by a moderate increase in the bandwidth (`pink beam' conditions) at some cost to data analysis complexity. Here, we report the first high-viscosity injector-based pink-beam SX experiments. The structures of proteinase K (PK) and A2A adenosine receptor (A2AAR) were determined to resolutions of 1.8 and 4.2 Å using 4 and 24 consecutive 100 ps X-ray pulse exposures, respectively. Strong PK data were processed using existing Laue approaches, while weaker A2AAR data required an alternative data-processing strategy. This demonstration of the feasibility presents new opportunities for time-resolved experiments with microcrystals to study structural changes in real time at pink-beam synchrotron beamlines worldwide.

Ultra-sensitive and selective determination of a phenolic food additive using protein capped gold nanoclusters: a dual in-line fluorometric and colorimetric sensing probe

2021 ◽  
Author(s):  
Sekar Shankar ◽  
N. S. K. Gowthaman ◽  
P. Arul ◽  
Feng Chen ◽  
H. N. Lim ◽  
...  
Keyword(s):  
Gold Nanoclusters ◽  
Food Additive ◽  
Colorimetric Sensing ◽  
Selective Determination ◽  
Red Luminescence ◽  
Colorimetric Methods

The application of red luminescence BSA-AuNCs towards the selective determination of food additive tert-butylhydroquinone (TBHQ) was demonstrated by both fluorometric and colorimetric methods.

Spherical SiO2 @ GdPO4 : Eu3+ Core–Shell Phosphors: Sol–Gel Synthesis and Characterization

2007 ◽  
Vol 7 (2) ◽  
pp. 542-548
Author(s):  
Cuikun Lin ◽  
Bo Zhao ◽  
Zhenling Wang ◽  
Min Yu ◽  
Huan Wang ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Low Voltage ◽  
Uv Light ◽  
Sol Gel ◽  
Emission Spectra ◽  
Core Shell ◽  
Time Resolved ◽  
Potential Applications ◽  
Ft Ir ◽  
Red Luminescence

Nanocrystalline GdPO4 : Eu3+ phosphor layers were coated on non-aggregated, monodisperse and spherical SiO2 particles by Pechini sol–gel method, resulting in the formation of core–shell structured SiO2 @ GdPO4 : Eu3+ particles. X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FT-IR), field emission scanning electron microscopy (FESEM), transmission electron microscopy (TEM), photoluminescence (PL), low-voltage cathodoluminescence (CL), time-resolved PL spectra and lifetimes were used to characterize the core–shell structured materials. Both XRD and FT-IR results indicate that GdPO4 layers have been successfully coated on the SiO2 particles, which can be further verified by the images of FESEM and TEM. Under UV light excitation, the SiO2 @ GdPO4 : Eu3+ phosphors show orange-red luminescence with Eu3+ 5D0–7F1 (593 nm) as the most prominent group. The PL excitation and emission spectra suggest that an energy transfer occurs from Gd3+ to Eu3+ in SiO2 @ GdPO4 : Eu3+ phosphors. The obtained core–shell phosphors have potential applications in FED and PDP devices.

Label-free fluorescence “turn-on” detection of SO32− by gold nanoclusters: integration in a hydrogel platform and intracellular detection

2019 ◽  
Vol 11 (9) ◽  
pp. 1214-1223 ◽  
Author(s):  
Abhay Sachdev ◽  
Rocky Raj ◽  
Ishita Matai ◽  
Vinay Kumar ◽  
P. Gopinath ◽  
...  
Keyword(s):  
Gold Nanoclusters ◽  
Label Free ◽  
Turn On ◽  
Potential Applications ◽  
Fluorescence Turn On ◽  
Intracellular Detection

A new dimension for the fluorescence “turn-on” detection of SO32− by Au NCs with potential applications towards the development of a hydrogel based platform and intracellular detection.

scholarly journals Au101-rGO nanocomposite: immobilization of phosphine-protected gold nanoclusters on reduced graphene oxide without aggregation

2021 ◽  
Author(s):  
Hanieh Mousavi ◽  
Yanting Yin ◽  
Liam Howard-Fabretto ◽  
Shailendra Kumar Sharma ◽  
Vladimir Golovko ◽  
...  
Keyword(s):  
Graphene Oxide ◽  
Transition Metal ◽  
Reduced Graphene Oxide ◽  
Metal Clusters ◽  
Gold Nanoclusters ◽  
Simple Approach ◽  
Transition Metal Clusters ◽  
Reduced Graphene ◽  
Potential Applications

Graphene supported transition metal clusters are of great interest for potential applications, such as catalysis, due to their unique properties. In this work, a simple approach to deposit Au101(PPh3)21Cl5 (Au101NC)...

scholarly journals Accurate Quantification of AAV Vector Genomes by Quantitative PCR

Genes ◽  
2021 ◽  
Vol 12 (4) ◽  
pp. 601
Author(s):  
Cristina Martinez-Fernandez de la Camara ◽  
Michelle McClements ◽  
Robert MacLaren
Keyword(s):  
Quantitative Pcr ◽  
Enzymatic Digestion ◽  
Absolute Quantification ◽  
Proteinase K ◽  
Aav Vector ◽  
Good Laboratory ◽  
Reference Plasmid ◽  
Pre Treatment ◽  
Dna Template ◽  
Accurate Quantification

The ability to accurately determine the dose of an adeno-associated viral (AAV) therapeutic vector is critical to the gene therapy process. Quantitative PCR (qPCR) is one of the common methods to quantify the AAV vector titre, but different variables can lead to inconsistent results. The aim of this study was to analyze the influence of the conformation of the DNA used as the standard control, and the enzymatic digestion was performed to release the viral genome from the protein capsid on the physical genome titration of a clinically relevant AAV8.RPGR vector, made to good laboratory practice standards in an academic setting. The results of this study showed that the conformation of the DNA used as standard has a significant impact on the accuracy of absolute quantification by qPCR. The use of supercoiled undigested plasmid DNA template generated a higher apparent titer, as compared to the use of linearized plasmid as the standard. In contrast to previous studies, the pre-treatment of the samples with Proteinase K, in addition to the high temperature step used after DNase I digestion, resulted in a reduction on AAV titers. Ideally, all AAV documentation should state which form of reference plasmid and which pre-treatment of the samples have been used to calculate titers, so that appropriate comparisons relating to dose toxicity and transduction efficacy can be made in the clinical scenario.

The contribution of the extracellular matrix to gravisensing in characean cells

1992 ◽  
Vol 101 (3) ◽  
pp. 611-623
Author(s):  
R. Wayne ◽  
M.P. Staves ◽  
A.C. Leopold
Keyword(s):  
Extracellular Matrix ◽  
Hydrolytic Enzymes ◽  
Chara Corallina ◽  
Proteinase K ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Carboxypeptidase B ◽  
Site Of Action ◽  
Protease Treatment ◽  
Essential Region

The cell-extracellular matrix junction, which includes the cell wall and the outer surface of the plasma membrane, may be an essential region for the perception of gravity by the internodal cells of Chara corallina. Typically, when an internodal cell is oriented vertically, the downwardly directed cytoplasmic stream travels at a velocity that is 10% faster than that of the upwardly directed stream. However when the cells are treated with impermeant hydrolytic enzymes that partially digest cellulose or hemicellulose, the cells lose their ability to respond to gravity even though streaming continues. By contrast, enzymes that digest pectins have no effect on the gravity-induced polarity of cytoplasmic streaming. Furthermore, gravisensing is sensitive to protease treatment; Proteinase K, thermolysin and collagenase but not trypsin, alpha-chymotrypsin or carboxypeptidase B, inhibit gravisensing. These findings indicate that proteins in the cell-extracellular matrix junction may be required for gravisensing. Moreover, the tetrapeptide Arg-Gly-Asp-Ser (RGDS) inhibits gravisensing in a concentration-dependent manner, indicating that the gravireceptor may be an integrin-like protein. The macromolecules necessary for gravisensing have been localized to the cell ends. As a consequence of the exoplasmic site of action of the enzymes and the tetrapeptides, we interpret the results to mean that they are acting on the gravireceptor, although we cannot eliminate the possibility that they are acting on the signal transduction chain. On the whole, our observations indicate that the cell-extracellular matrix junction is a sine qua non for graviperception in statolith-free Chara internodal cells and we suggest that the gravireceptor is located in this region.

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