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Theory of Polymer Entrapped Enzyme Ultramicroelectrodes: Application to Glucose and Adenosine Triphosphate Detection.

ABSTRACT: We validate, by comparison with experimental data, a theoretical description of the amperometric response of microbiosensors formed via enzyme entrapment. The utility of the theory is further illustrated with two relevant examples supported by experiments: (1) quantitative detection of glucose and (2) quantitative detection of adenosine triphosphate (ATP).

SUBMITTER: Kottke PA

PROVIDER: S-EPMC2863125 | biostudies-literature | 2008 Jul

REPOSITORIES: biostudies-literature

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Theory of Polymer Entrapped Enzyme Ultramicroelectrodes: Application to Glucose and Adenosine Triphosphate Detection.

Kottke Peter A PA Kranz Christine C Kwon Yong Koo YK Masson Jean-Francois JF Mizaikoff Boris B Fedorov Andrei G AG

Journal of electroanalytical chemistry (Lausanne, Switzerland) 20080701 1-2

We validate, by comparison with experimental data, a theoretical description of the amperometric response of microbiosensors formed via enzyme entrapment. The utility of the theory is further illustrated with two relevant examples supported by experiments: (1) quantitative detection of glucose and (2) quantitative detection of adenosine triphosphate (ATP). ...[more]

PMID: 20445817

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Theory of Polymer Entrapped Enzyme Ultramicroelectrodes: Fundamentals.

Project description:We have developed a theoretical description of the amperometric response of ultramicroelectrode (UME) biosensors formed via enzyme entrapment. Our model allows for multiple enzymes and co-substrates, and results in a closed-form analytical expression for the steady-state current response of the disk ultramicroelectrode. It captures the effects of enzyme-entrapment domain size, species transport properties (which can be different in the polymer and surrounding electrolyte), enzyme kinetics, and axisymmetric diffusion. Assumptions inherent in the derivation are carefully explained, as are the resulting limits on the applicability of the results. The ability to theoretically predict the response of enzyme entrapped UMEs should enable improved design, operation, and data interpretation for this important class of biosensors.

| S-EPMC2863126 | biostudies-literature

Simultaneous Detection of Adenosine Triphosphate and Glucose Based on the Cu-Fenton Reaction.

Project description:Both adenosine triphosphate (ATP) and glucose are important to human health, and their abnormal levels are closely related to angiocardiopathy and hypoglycaemia. Therefore, the simultaneous determination of ATP and glucose with a single test mode is highly desirable for disease diagnostics and early recognition. Herein, a new fluorescence on/off switch sensing platform is developed by carbon nanodots (CNDs) to detect ATP and glucose simultaneously. The fluorescence of CNDs can be quenched by Cu2+ and hydrogen peroxide (H?O?), due to the formation of hydroxyl radicals (·OH) produced in the Cu-Fenton reaction. Based on the high affinity of Cu2+ with ATP, the fluorescence of CNDs will recover effectively after adding ATP. Additionally, glucose can be efficiently catalyzed by glucose oxidase (GOx) to generate H?O?, so the platform can also be utilized to analyze glucose. Under optimum conditions, this sensing platform displays excellent sensitivity and the linear ranges are from 0.1 to 7 μM for ATP with a limit of detection (LOD) of 30.2 nM, and from 0.1 to 7 mM for glucose with a LOD 39.8 μM, respectively. Benefiting from the high sensitivity and selectivity, this sensing platform is successfully applied for simultaneous detection of ATP and glucose in human serum samples with satisfactory recoveries.

| S-EPMC6069456 | biostudies-literature

Ex vivo and in vivo fluorescence detection and imaging of adenosine triphosphate.

Project description:BackgroundEx vivo and in vivo detection and imaging of adenosine triphosphate (ATP) is critically important for the diagnosis and treatment of diseases, which still remains challenges up to present.ResultsWe herein demonstrate that ATP could be fluorescently detected and imaged ex vivo and in vivo. In particular, we fabricate a kind of fluorescent ATP probes, which are made of titanium carbide (TC) nanosheets modified with the ROX-tagged ATP-aptamer (TC/Apt). In the constructed TC/Apt, TC shows superior quenching efficiency against ROX (e.g., ~ 97%). While in the presence of ATP, ROX-tagged aptamer is released from TC surface, leading to the recovery of fluorescence of ROX under the 545-nm excitation. Consequently, a wide dynamic range from 1 μM to 1.5 mM ATP and a high sensitivity with a limit of detection (LOD) down to 0.2 μM ATP can be readily achieved by the prepared TC/Apt. We further demonstrate that the as-prepared TC/Apt probe is feasible for accurate discrimination of ATP in different samples including living cells, body fluids (e.g., mouse serum, mouse urine and human serum) and mouse tumor models.ConclusionsFluorescence detection and imaging of ATP could be readily achieved in living cells, body fluids (e.g., urine and serum), as well as mouse tumor model through a new kind of fluorescent ATP nanoprobes, offering new powerful tools for the treatment of diseases related to abnormal fluctuation of ATP concentration.

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CRISPR/Cas-Assisted Nanoneedle Sensor for Adenosine Triphosphate Detection in Living Cells.

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Transition State Analysis of Adenosine Triphosphate Phosphoribosyltransferase.

Project description:Adenosine triphosphate phosphoribosyltransferase (ATP-PRT) catalyzes the first step in histidine biosynthesis, a pathway essential to microorganisms and a validated target for antimicrobial drug design. The ATP-PRT enzyme catalyzes the reversible substitution reaction between phosphoribosyl pyrophosphate and ATP. The enzyme exists in two structurally distinct forms, a short- and a long-form enzyme. These forms share a catalytic core dimer but bear completely different allosteric domains and thus distinct quaternary assemblies. Understanding enzymatic transition states can provide essential information on the reaction mechanisms and insight into how differences in domain structure influence the reaction chemistry, as well as providing a template for inhibitor design. In this study, the transition state structures for ATP-PRT enzymes from Campylobacter jejuni and Mycobacterium tuberculosis (long-form enzymes) and from Lactococcus lactis (short-form) were determined and compared. Intrinsic kinetic isotope effects (KIEs) were obtained at reaction sensitive positions for the reverse reaction using phosphonoacetic acid, an alternative substrate to the natural substrate pyrophosphate. The experimental KIEs demonstrated mechanistic similarities between the three enzymes and provided experimental boundaries for quantum chemical calculations to characterize the transition states. Predicted transition state structures support a dissociative reaction mechanism with a DN*AN‡ transition state. Weak interactions from the incoming nucleophile and a fully dissociated ATP adenine are predicted regardless of the difference in overall structure and quaternary assembly. These studies establish that despite significant differences in the quaternary assembly and regulatory machinery between ATP-PRT enzymes from different sources, the reaction chemistry and catalytic mechanism are conserved.

| S-EPMC6693643 | biostudies-literature

Some properties of rat-liver glucose--adenosine triphosphate phosphotransferases.

Project description:In normal rat liver hexokinase (EC 2.7.1.1) activity usually accounts for not more than 30% of the total glucose-ATP phosphotransferase activity, the remainder being due to glucokinase (EC 2.7.1.2). In the present work it was found that in normal rat liver the relative activities of these two enzymes were occasionally very different from those usually found even though the total glucose-ATP phosphotransferase activity was within the normal range. In some cases almost the entire glucose-ATP phosphotransferase was accounted for by the low-K(m) enzyme hexokinase. Some properties of this enzyme system are reported. It is suggested that this shift in favour of the low-K(m) enzyme without change in the total glucose-ATP phosphotransferase activity may represent a regulatory mechanism.

| S-EPMC1265217 | biostudies-other

Adenosine triphosphate stimulates Aquifex aeolicus MutL endonuclease activity.

Project description:BackgroundHuman PMS2 (hPMS2) homologues act to nick 5' and 3' to misincorporated nucleotides during mismatch repair in organisms that lack MutH. Mn(++) was previously found to stimulate the endonuclease activity of these homologues. ATP was required for the nicking activity of hPMS2 and yPMS1, but was reported to inhibit bacterial MutL proteins from Thermus thermophilus and Aquifex aeolicus that displayed homology to hPMS2. Mutational analysis has identified the DQHA(X)(2)E(X)(4)E motif present in the C-terminus of PMS2 homologues as important for endonuclease activity.Methodologies/principal findingsWe examined the effect ATP had on the Mn(++) induced nicking of supercoiled pBR322 by full-length and mutant A. aeolicus MutL (Aae MutL) proteins. Assays were single time point, enzyme titration experiments or reaction time courses. The maximum velocity for MutL nicking was determined to be 1.6+/-0.08x10(-5) s(-1) and 4.2+/-0.3x10(-5) s(-1) in the absence and presence of ATP, respectively. AMPPNP stimulated the nicking activity to a similar extent as ATP. A truncated Aae MutL protein composed of only the C-terminal 123 amino acid residues was found to nick supercoiled DNA. Furthermore, mutations in the conserved C-terminal DQHA(X)(2)E(X)(4)E and CPHGRP motifs were shown to abolish Aae MutL endonuclease activity.ConclusionsATP stimulated the Mn(++) induced endonuclease activity of Aae MutL. Experiments utilizing AMPPNP implied that the stimulation did not require ATP hydrolysis. A mutation in the DQHA(X)(2)E(X)(4)E motif of Aae MutL further supported the role of this region in endonclease activity. For the first time, to our knowledge, we demonstrate that changing the histidine residue in the conserved CPHGRP motif abolishes endonucleolytic activity of a hPMS2 homologue. Finally, the C-terminal 123 amino acid residues of Aae MutL were sufficient to display Mn(++) induced nicking activity.

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A calcium ion-dependent adenosine triphosphate pyrophosphohydrolase in plasma membrane from rat liver. Demonstration that the adenosine triphosphate analogues adenosine 5'-[betagamma-imido]triphosphate and adenosine 5'-[betagamma-methylene]-triphosphate are substrates for the enzyme.

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Overcoming Debye length limitations: Three-dimensional wrinkled graphene field-effect transistor for ultra-sensitive adenosine triphosphate detection.

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| S-EPMC10205565 | biostudies-literature

Synthesis of a Biologically Important Adenosine Triphosphate Analogue, ApppD.

Project description:The chemical synthesis of a adenosine triphosphate analogue, 1-adenosin-5'-yl 3-(3-methylbut-2-enyl) triphosphoric acid diester (ApppD), is described. ApppD is known to be an active metabolite of the mevalonate pathway in the human body like its structural isomer isopentenyl ester of ATP (ApppI). Very recently, ApppI has been found to possess novel function(s); now it will also be possible to examine the effects of ApppD more precisely because it can be synthesized in reasonable amounts. 1-Adenosin-5'-yl 3-(3-methylbut-2-enyl) diphosphoric acid diester (AppD; a adenosine diphosphate analogue) was also isolated from the synthesis mixture. Both ApppD and AppD were characterized by 1H, 13C, 31P NMR and mass spectrometry methods.

| S-EPMC6044877 | biostudies-literature

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