Browse
Submit Data
Databases
API
Help

Dataset Information

22 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Synthetic biosensors for precise gene control and real-time monitoring of metabolites.

ABSTRACT: Characterization and standardization of inducible transcriptional regulators has transformed how scientists approach biology by allowing precise and tunable control of gene expression. Despite their utility, only a handful of well-characterized regulators exist, limiting the complexity of engineered biological systems. We apply a characterization pipeline to four genetically encoded sensors that respond to acrylate, glucarate, erythromycin and naringenin. We evaluate how the concentration of the inducing chemical relates to protein expression, how the extent of induction affects protein expression kinetics, and how the activation behavior of single cells relates to ensemble measurements. We show that activation of each sensor is orthogonal to the other sensors, and to other common inducible systems. We demonstrate independent control of three fluorescent proteins in a single cell, chemically defining eight unique transcriptional states. To demonstrate biosensor utility in metabolic engineering, we apply the glucarate biosensor to monitor product formation in a heterologous glucarate biosynthesis pathway and identify superior enzyme variants. Doubling the number of well-characterized inducible systems makes more complex synthetic biological circuits accessible. Characterizing sensors that transduce the intracellular concentration of valuable metabolites into fluorescent readouts enables high-throughput screening of biological catalysts and alleviates the primary bottleneck of the metabolic engineering design-build-test cycle.

SUBMITTER: Rogers JK

PROVIDER: S-EPMC4551912 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Cell membrane-anchored biosensors for real-time monitoring of the cellular microenvironment.

Project description:Cell membrane-anchored biochemical sensors that allow real-time monitoring of the interactions of cells with their microenvironment would be powerful tools for studying the mechanisms underlying various biological processes, such as cell metabolism and signaling. Despite the significance of these techniques, unfortunately, their development has lagged far behind due to the lack of a desirable membrane engineering method. Here, we propose a simple, efficient, biocompatible, and universal strategy for one-step self-construction of cell-surface sensors using diacyllipid-DNA conjugates as the building and sensing elements. The sensors exploit the high membrane-insertion capacity of a diacyllipid tail and good sensing performance of the DNA probes. Based on this strategy, we have engineered specific DNAzymes on the cell membrane for metal ion assay in the extracellular microspace. The immobilized DNAzyme showed excellent performance for reporting and semiquantifying both exogenous and cell-extruded target metal ions in real time. This membrane-anchored sensor could also be used for multiple target detection by having different DNA probes inserted, providing potentially useful tools for versatile applications in cell biology, biomedical research, drug discovery, and tissue engineering.

| S-EPMC4183634 | biostudies-literature

Real-time monitoring biomarker expression of carcinoma cells by surface plasmon resonance biosensors.

Project description:A novel surface plasmon resonance (SPR) biosensor which is capable of monitoring proteomic biomarker secretion from living cells is reported here. Vascular endothelial growth factor (VEGF) secretion from living SKOV-3 ovarian cancer cells was measured for concept demonstration.

| S-EPMC3496773 | biostudies-literature

Monitoring Proteolytic Activity in Real Time: A New World of Opportunities for Biosensors.

Project description:Proteases play a pivotal role in several biological processes, from digestion, cell proliferation, and differentiation to fertility. Deregulation of protease metabolism can result in several pathological conditions (i.e., cancer, neurodegenerative disorders, and others). Therefore, monitoring proteolytic activity in real time could have a fundamental role in the early diagnosis of these diseases. Herein, the main approaches used to develop biosensors for monitoring proteolytic activity are reviewed. A comparison of the advantages and disadvantages of each approach is provided along with a discussion of their importance and promising opportunities for the early diagnosis of severe diseases. This new era of biosensors can be characterized by the ability to control and monitor biological processes, ultimately improving the potential of personalized medicine.

| S-EPMC7199732 | biostudies-literature

DNA-SWCNT Biosensors Allow Real-Time Monitoring of Therapeutic Responses in Pancreatic Ductal Adenocarcinoma.

Project description:Pancreatic ductal adenocarcinoma (PDAC) is a highly desmoplastic cancer with limited treatment options. There is an urgent need for tools that monitor therapeutic responses in real time. Drugs such as gemcitabine and irinotecan elicit their therapeutic effect in cancer cells by producing hydrogen peroxide (H2O2). In this study, specific DNA-wrapped single-walled carbon nanotubes (SWCNT), which precisely monitor H2O2, were used to determine the therapeutic response of PDAC cells in vitro and tumors in vivo. Drug therapeutic efficacy was evaluated in vitro by monitoring H2O2 differences in situ using reversible alteration of Raman G-bands from the nanotubes. Implantation of the DNA-SWCNT probe inside the PDAC tumor resulted in approximately 50% reduction of Raman G-band intensity when treated with gemcitabine versus the pretreated tumor; the Raman G-band intensity reversed to its pretreatment level upon treatment withdrawal. In summary, using highly specific and sensitive DNA-SWCNT nanosensors, which can determine dynamic alteration of hydrogen peroxide in tumor, can evaluate the effectiveness of chemotherapeutics. SIGNIFICANCE: A novel biosensor is used to detect intratumoral hydrogen peroxide, allowing real-time monitoring of responses to chemotherapeutic drugs.

| S-EPMC6726513 | biostudies-literature

Plasmonic biosensors fabricated by galvanic displacement reactions for monitoring biomolecular interactions in real time.

Project description:Optical sensors are prepared by reduction of gold ions using freshly etched hydride-terminated porous silicon, and their ability to specifically detect binding between protein A/rabbit IgG and asialofetuin/Erythrina cristagalli lectin is studied. The fabrication process is simple, fast, and reproducible, and does not require complicated lab equipment. The resulting nanostructured gold layer on silicon shows an optical response in the visible range based on the excitation of localized surface plasmon resonance. Variations in the refractive index of the surrounding medium result in a color change of the sensor which can be observed by the naked eye. By monitoring the spectral position of the localized surface plasmon resonance using reflectance spectroscopy, a bulk sensitivity of 296 nm?±?3 nm/RIU is determined. Furthermore, selectivity to target analytes is conferred to the sensor through functionalization of its surface with appropriate capture probes. For this purpose, biomolecules are deposited either by physical adsorption or by covalent coupling. Both strategies are successfully tested, i.e., the optical response of the sensor is dependent on the concentration of respective target analyte in the solution facilitating the determination of equilibrium dissociation constants for protein A/rabbit IgG as well as asialofetuin/Erythrina cristagalli lectin which are in accordance with reported values in literature. These results demonstrate the potential of the developed optical sensor for cost-efficient biosensor applications. Graphical abstract.

| S-EPMC7214386 | biostudies-literature

Synthetic-type control charts for time-between-events monitoring.

Project description:This paper proposes three synthetic-type control charts to monitor the mean time-between-events of a homogenous Poisson process. The first proposed chart combines an Erlang (cumulative time between events, Tr ) chart and a conforming run length (CRL) chart, denoted as Synth-Tr chart. The second proposed chart combines an exponential (or T) chart and a group conforming run length (GCRL) chart, denoted as GR-T chart. The third proposed chart combines an Erlang chart and a GCRL chart, denoted as GR-Tr chart. By using a Markov chain approach, the zero- and steady-state average number of observations to signal (ANOS) of the proposed charts are obtained, in order to evaluate the performance of the three charts. The optimal design of the proposed charts is shown in this paper. The proposed charts are superior to the existing T chart, Tr chart, and Synth-T chart. As compared to the EWMA-T chart, the GR-T chart performs better in detecting large shifts, in terms of the zero- and steady-state performances. The zero-state Synth-T4 and GR-Tr (r = 3 or 4) charts outperform the EWMA-T chart for all shifts, whereas the Synth-Tr (r = 2 or 3) and GR-T 2 charts perform better for moderate to large shifts. For the steady-state process, the Synth-Tr and GR-Tr charts are more efficient than the EWMA-T chart in detecting small to moderate shifts.

| S-EPMC3670912 | biostudies-literature

In-vivo real-time control of protein expression from endogenous and synthetic gene networks.

Project description:We describe an innovative experimental and computational approach to control the expression of a protein in a population of yeast cells. We designed a simple control algorithm to automatically regulate the administration of inducer molecules to the cells by comparing the actual protein expression level in the cell population with the desired expression level. We then built an automated platform based on a microfluidic device, a time-lapse microscopy apparatus, and a set of motorized syringes, all controlled by a computer. We tested the platform to force yeast cells to express a desired fixed, or time-varying, amount of a reporter protein over thousands of minutes. The computer automatically switched the type of sugar administered to the cells, its concentration and its duration, according to the control algorithm. Our approach can be used to control expression of any protein, fused to a fluorescent reporter, provided that an external molecule known to (indirectly) affect its promoter activity is available.

| S-EPMC4022480 | biostudies-literature

Real-Time Monitoring of the Yeast Intracellular State During Bioprocesses With a Toolbox of Biosensors.

Project description:Industrial fermentation processes strive for high robustness to ensure optimal and consistent performance. Medium components, fermentation products, and physical perturbations may cause stress and lower performance. Cellular stress elicits a range of responses, whose extracellular manifestations have been extensively studied; whereas intracellular aspects remain poorly known due to lack of tools for real-time monitoring. Genetically encoded biosensors have emerged as promising tools and have been used to improve microbial productivity and tolerance toward industrially relevant stresses. Here, fluorescent biosensors able to sense the yeast intracellular environment (pH, ATP levels, oxidative stress, glycolytic flux, and ribosome production) were implemented into a versatile and easy-to-use toolbox. Marker-free and efficient genome integration at a conserved site on chromosome X of Saccharomyces cerevisiae strains and a commercial Saccharomyces boulardii strain was developed. Moreover, multiple biosensors were used to simultaneously monitor different intracellular parameters in a single cell. Even when combined together, the biosensors did not significantly affect key physiological parameters, such as specific growth rate and product yields. Activation and response of each biosensor and their interconnection were assessed using an advanced micro-cultivation system. Finally, the toolbox was used to screen cell behavior in a synthetic lignocellulosic hydrolysate that mimicked harsh industrial substrates, revealing differences in the oxidative stress response between laboratory (CEN.PK113-7D) and industrial (Ethanol Red) S. cerevisiae strains. In summary, the toolbox will allow both the exploration of yeast diversity and physiological responses in natural and complex industrial conditions, as well as the possibility to monitor production processes.

| S-EPMC8776715 | biostudies-literature

Precise Spatially Selective Photothermolysis Using Modulated Femtosecond Lasers and Real-time Multimodal Microscopy Monitoring.

Project description:The successful application of lasers in the treatment of skin diseases and cosmetic surgery is largely based on the principle of conventional selective photothermolysis which relies strongly on the difference in the absorption between the therapeutic target and its surroundings. However, when the differentiation in absorption is not sufficient, collateral damage would occur due to indiscriminate and nonspecific tissue heating. To deal with such cases, we introduce a novel spatially selective photothermolysis method based on multiphoton absorption in which the radiant energy of a tightly focused near-infrared femtosecond laser beam can be directed spatially by aiming the laser focal point to the target of interest. We construct a multimodal optical microscope to perform and monitor the spatially selective photothermolysis. We demonstrate that precise alteration of the targeted tissue is achieved while leaving surrounding tissue intact by choosing appropriate femtosecond laser exposure with multimodal optical microscopy monitoring in real time.

| S-EPMC5327629 | biostudies-literature

Ex vivo real-time monitoring of volatile metabolites resulting from nasal odorant metabolism.

Project description:Odorant-metabolizing enzymes are critically involved in the clearance of odorant molecules from the environment of the nasal neuro-olfactory tissue to maintain the sensitivity of olfactory detection. Odorant metabolism may also generate metabolites in situ, the characterization and function of which in olfaction remain largely unknown. Here, we engineered and validated an ex vivo method to measure odorant metabolism in real-time. Glassware containing an explant of rat olfactory mucosa was continuously flushed with an odorant flow and was coupled to a proton transfer reaction-mass spectrometer for volatile compound analysis. Focusing on carboxylic esters and diketone odorants, we recorded the metabolic uptake of odorants by the mucosa, concomitantly with the release of volatile odorant metabolites in the headspace. These results significantly change the picture of real-time in situ odorant metabolism and represent a new step forward in the investigation of the function of odorant metabolites in the peripheral olfactory process. Our method allows the systematic identification of odorant metabolites using a validated animal model and permits the screening of olfactory endogenously produced chemosensory molecules.

| S-EPMC6385289 | biostudies-other

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

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