Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Covalent Attachment of Active Enzymes to Upconversion Phosphors Allows Ratiometric Detection of Substrates.

ABSTRACT: Upconverting phosphors (UCPs) convert multiple low energy photons into higher energy emission via the process of photon upconversion and offer an attractive alternative to organic fluorophores for use as luminescent probes. Here, UCPs were capped with functionalized silica in order to provide a surface to covalently conjugate proteins with surface-accessible cysteines. Variants of green fluorescent protein (GFP) and the flavoenzyme pentaerythritol tetranitrate reductase (PETNR) were then attached via maleimide-thiol coupling in order to allow energy transfer from the UCP to the GFP or flavin cofactor of PETNR, respectively. PETNR retains its activity when coupled to the UCPs, which allows reversible detection of enzyme substrates via ratiometric sensing of the enzyme redox state.

SUBMITTER: Burgess L

PROVIDER: S-EPMC7756657 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Covalent Attachment of Active Enzymes to Upconversion Phosphors Allows Ratiometric Detection of Substrates.

Burgess Letitia L Wilson Hannah H Jones Alex R AR Harvey Peter P Natrajan Louise S LS Hay Sam S

Chemistry (Weinheim an der Bergstrasse, Germany) 20201016 65

Upconverting phosphors (UCPs) convert multiple low energy photons into higher energy emission via the process of photon upconversion and offer an attractive alternative to organic fluorophores for use as luminescent probes. Here, UCPs were capped with functionalized silica in order to provide a surface to covalently conjugate proteins with surface-accessible cysteines. Variants of green fluorescent protein (GFP) and the flavoenzyme pentaerythritol tetranitrate reductase (PETNR) were then attache ...[more]

PMID: 32476171

Similar Datasets

Assessing the Covalent Attachment and Energy Transfer Capabilities of Upconverting Phosphors With Cofactor Containing Bioactive Enzymes.

Project description:Upconverting phosphors (UCPs) convert multiple low energy photons into higher energy emission via the process of photon upconversion and offer an attractive alternative to organic fluorophores for use as luminescent probes. Examples of biosensors utilizing the apparent energy transfer of UCPs and nanophosphors (UCNPs) with biomolecules have started to appear in the literature but very few exploit the covalent anchoring of the biomolecule to the surface of the UCP to improve the sensitivity of the systems. Here, we demonstrate a robust and versatile method for the covalent attachment of biomolecules to the surface of a variety of UCPs and UCNPs in which the UCPs were capped with functionalized silica in order to provide a surface to covalently conjugate biomolecules with surface-accessible cysteines. Variants of BM3Heme, cytochrome C, glucose oxidase, and glutathione reductase were then attached via maleimide-thiol coupling. BM3Heme, glucose oxidase, and glutathione reductase were shown to retain their activity when coupled to the UCPs potentially opening up opportunities for biosensing applications.

| S-EPMC7779683 | biostudies-literature

A Strategy of NIR Dual-Excitation Upconversion for Ratiometric Intracellular Detection.

Project description:Intracellular detection is highly desirable for biological research and clinical diagnosis, yet its quantitative analysis with noninvasivity, sensitivity, and accuracy remains challenging. Herein, a near-infrared (NIR) dual-excitation strategy is reported for ratiometric intracellular detection through the design of dye-sensitized upconversion probes and employment of a purpose-built NIR dual-laser confocal microscope. NIR dye IR808, a recognizer of intracellular analyte hypochlorite, is introduced as energy donor and Yb,Er-doped NaGdF4 upconversion nanoparticles are adopted as energy acceptor in the as-designed nanoprobes. The efficient analyte-dependent energy transfer and low background luminescence endow the nanoprobes with ultrahigh sensitivity. In addition, with the nonanalyte-dependent upconversion luminescence (UCL) excited by 980 nm as a self-calibrated signal, the interference from environmental fluctuation can be alleviated. Furthermore, the dual 808/980 nm excited ratiometric UCL is demonstrated for the quantification of the level of intracellular hypochlorite. Particularly, the intrinsic hypochlorite with only nanomolar concentration in live MCF-7 cells in the absence of exogenous stimuli is determined. Such an NIR dual-excitation ratiometric strategy based on dye-sensitized UCL probes can be easily extended to detect various intracellular analytes through tailoring the reactive NIR dyes, which provides a promising tool for probing biochemical processes in live cells and diagnosing diseases.

| S-EPMC6864516 | biostudies-literature

Covalent Attachment of Enzymes to Paper Fibers for Paper-Based Analytical Devices.

Project description:Due to its unique material properties, paper offers many practical advantages as a viable platform for sensing devices. In view of paper-based microfluidic biosensing applications, the covalent immobilization of enzymes with preserved functional activity is highly desirable and ultimately challenging. In the present manuscript, we report an efficient approach to achieving the covalent attachment of certain enzymes on paper fibers via a surface-bound network of hydrophilic polymers bearing protein-modifiable sites. This tailor-made macromolecular system consisting of polar, highly swellable copolymers is anchored to the paper exterior upon light-induced crosslinking of engineered benzophenone motifs. On the other hand, this framework contains active esters that can be efficiently modified by the nucleophiles of biomolecules. This strategy allowed the covalent immobilization of glucose oxidase and horseradish peroxidase onto cotton linters without sacrificing their bioactivities and performance upon surface binding. As a proof-of-concept application, a microfluidic chromatic paper-based glucose sensor was developed and achieved successful glucose detection in a simple yet efficient cascade reaction.

| S-EPMC6030327 | biostudies-literature

A hybrid upconversion nanoprobe for ratiometric detection of aliphatic biogenic amines in aqueous medium.

Project description:We fabricated an inorganic-organic hybrid upconversion nanoprobe for the ratiometric detection of aliphatic biogenic amines in water. The hybrid nanoprobe comprises a thiophene-based acceptor-π-donor-π-acceptor organic fluorescent dye, TDPM, and near-infrared light-absorbing upconversion nanoparticles (UCNPs). The organic dye was loaded into a mesoporous silica-coated UCNP (UCNP@mSiO2) matrix to circumvent the issues of water insolubility and higher energy excitation. Yb3+ and Tm3+-doped UCNPs exhibited dual emission bands at 475 and 645 nm upon excitation with a 980 nm laser. The significant spectral overlap between the absorption and the emission bands of TDPM and UCNPs, respectively, at 475 nm led to resonance energy transfer (RET) from the UCNPs to TDPM resulting in the quenching of the UCNP emission. In contrast, 'turn-on' emission was noticeable with the addition of aliphatic biogenic amines due to an inhibition of the RET. The emission at 645 nm remained unaffected during the energy transfer process making the hybrid probe a versatile platform for the ratiometric detection of different aliphatic biogenic amines. Furthermore, we explored the sensing of aliphatic biogenic amines in adulterated milk and rotten fish. The unique material attributes demonstrated in the current study hold promise for further development of real-time sensors and switches based on hybrid upconversion nanoprobes.

| S-EPMC9419630 | biostudies-literature

Rationally designed upconversion nanoprobe for simultaneous highly sensitive ratiometric detection of fluoride ions and fluorosis theranostics.

Project description:For many years, fluorosis has been known as a worldwide disease which seriously diminishes quality of life through skeletal embrittlement and hepatic damage. Aiming to develop novel drugs for simultaneous fluorosis diagnosis and therapy, in this work we explore the feasibility of a novel pyrogallic acid-titanium(iv) complex-modified upconversion nanoprobe (UCNP-PA-Ti) for F- capture and real-time quantification. Utilizing the strong interaction between Ti4+ and F-, the modified PA-Ti decomposes in F--containing solution, which not only weakens the FRET but results in upconversion luminescence (UCL) recovery. Both in vitro and in vivo experiments demonstrate a highly sensitive F- UCL response and therapeutic efficiency, which was promising for successful UCL image monitoring and the therapeutic process. Long blood circulation time and low toxicity ensured their safe application for fluorosis theranostics. Our work provides a new possibility for F- concentration detection within fluorosis therapeutic periods and encourages the development of novel drugs for fluorosis theranostics.

| S-EPMC6001250 | biostudies-literature

Covalent attachment of lipid vesicles to a fluid-supported bilayer allows observation of DNA-mediated vesicle interactions.

Project description:Specific membrane interactions such as lipid vesicle docking and fusion can be mediated by synthetic DNA-lipid conjugates as a model for the protein-driven processes that are ubiquitous in biological systems. Here we present a method of tethering vesicles to a supported lipid bilayer that allows the simultaneous deposition of cognate vesicle partners displaying complementary DNA, resulting in well-mixed populations of tethered vesicles that are laterally mobile. Vesicles are covalently attached to a supporting lipid bilayer using a DNA-templated click reaction; then DNA-mediated interactions between tethered vesicles are triggered by spiking the salt concentration. These interactions, such as docking and fusion, can then be observed for individual vesicles as they collide on the surface. The architecture of this new system also permits control over the number of lipid anchors that tether the vesicle to the supporting bilayer. The diffusion coefficient of tethered vesicles anchored by two lipids is approximately 1.6-fold slower than that of vesicles anchored only with a single lipid, consistent with a simple physical model.

| S-EPMC2877162 | biostudies-literature

Ratiometric nanothermometer in vivo based on triplet sensitized upconversion.

Project description:Temperature is an essential factor that counts for living systems where complicated vital activities are usually temperature dependent. In vivo temperature mapping based on non-contact optical approach will be beneficial for revealing the physiological phenomena behind with minimized influence to the organism. Herein, a highly thermal-sensitive upconversion system based on triplet-triplet annihilation (TTA) mechanism is pioneered to indicate body temperature variation sensitively over the physiological temperature range. The temperature-insensitive NaYF4: Nd nanophosphors with NIR emission was incorporated into the temperature-responsive TTA-upconversion system to serve as an internal calibration unit. Consequently, a ratiometric thermometer capable of accurately monitoring the temperature changes in vivo was developed with high thermal sensitivity (~7.1% K-1) and resolution (~0.1?K).

| S-EPMC6043590 | biostudies-literature

Highly Selective and Sensitive Ratiometric Detection of Sn2+ Ions Using NIR-Excited Rhodamine-B-Linked Upconversion Nanophosphors

Project description:Detection of Sn2+ ions in environmental and

| S-EPMC9434793 | biostudies-literature

Lanthanide-doped bismuth-based nanophosphors for ratiometric upconversion optical thermometry.

Project description:Nanothermometry could realize stable, efficient, and noninvasive temperature detection at the nanoscale. Unfortunately, most applications of nanothermometers are still limited due to their intricate synthetic process and low-temperature sensitivity. Herein, we reported a kind of novel bismuth-based upconversion nanomaterial with a fast and facile preparation strategy. The bismuth-based upconversion luminophore was synthesized by the co-precipitation method within 1 minute. By optimizing the doping ratio of the sensitizer Yb ion and the activator Er ion and adjusting the synthetic solvent strategy, the crystallinity of the nanomaterials was increased and the upconversion luminescence intensity was improved. Ratiometric upconversion optical measurements of temperature in the range of 278 K to 358 K can be achieved by ratiometric characteristic emission peaks of thermally sensitive Er ion. This method of rapidly constructing nanometer temperature probes provides a feasible strategy for the construction of novel fluorescent temperature probes.

| S-EPMC8985227 | biostudies-literature

Predicting novel substrates for enzymes with minimal experimental effort with active learning.

Project description:Enzymatic substrate promiscuity is more ubiquitous than previously thought, with significant consequences for understanding metabolism and its application to biocatalysis. This realization has given rise to the need for efficient characterization of enzyme promiscuity. Enzyme promiscuity is currently characterized with a limited number of human-selected compounds that may not be representative of the enzyme's versatility. While testing large numbers of compounds may be impractical, computational approaches can exploit existing data to determine the most informative substrates to test next, thereby more thoroughly exploring an enzyme's versatility. To demonstrate this, we used existing studies and tested compounds for four different enzymes, developed support vector machine (SVM) models using these datasets, and selected additional compounds for experiments using an active learning approach. SVMs trained on a chemically diverse set of compounds were discovered to achieve maximum accuracies of ~80% using ~33% fewer compounds than datasets based on all compounds tested in existing studies. Active learning-selected compounds for testing resolved apparent conflicts in the existing training data, while adding diversity to the dataset. The application of these algorithms to wide arrays of metabolic enzymes would result in a library of SVMs that can predict high-probability promiscuous enzymatic reactions and could prove a valuable resource for the design of novel metabolic pathways.

| S-EPMC7055960 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data