Glucose sensor for low-cost lifetime-based sensing using a genetically engineered protein.

L Tolosa; I Gryczynski; L R Eichhorn; J D Dattelbaum; F N Castellano; G Rao; J R Lakowicz

doi:10.1006/abio.1998.2974

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PubMed · 9918662

Glucose sensor for low-cost lifetime-based sensing using a genetically engineered protein.

L Tolosa·I Gryczynski·L R Eichhorn·J D Dattelbaum·F N Castellano·G Rao·J R Lakowicz

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Abstract

We describe a glucose sensor based on a mutant glucose/galactose binding protein (GGBP) and phase-modulation fluorometry. The GGBP from Escherichia coli was mutated to contain a single cysteine residue at position 26. When labeled with a sulfhydryl-reactive probe 2-(4'-iodoacetamidoanilino)naphthalene-6-sulfonic acid, the labeled protein displayed a twofold decrease in intensity in response to glucose, with a dissociation constant near 1 microM glucose. The ANS-labeled protein displayed only a modest change in lifetime, precluding lifetime-based sensing of glucose. A modulation sensor was created by combining ANS26-GGBP with a long-lifetime ruthenium (Ru) metal-ligand complex on the surface of the cuvette. Binding of glucose changed the relative intensity of ANS26-GGBP and the Ru complex, resulting in a dramatic change in modulation at a low frequency of 2.1 MHz. Modulation measurements at 2.1 MHz were shown to accurately determine the glucose concentration. These results suggest an approach to glucose sensing with simple devices.

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Bacterial Proteins Biosensing Techniques Costs and Cost Analysis Escherichia coli Fluorescent Dyes Glucose Models, Molecular Monosaccharide Transport Proteins Mutagenesis, Site-Directed Naphthalenesulfonates Protein Conformation Protein Engineering Recombinant Proteins Spectrometry, Fluorescence Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

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L Tolosa, I Gryczynski, L R Eichhorn, J D Dattelbaum, F N Castellano, G Rao, J R Lakowicz. 1999-02-01. Glucose sensor for low-cost lifetime-based sensing using a genetically engineered protein.. https://doi.org/10.1006/abio.1998.2974

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