A modified fluorescent sensor for reporting glucose concentration in the airway lumen

• Published in: PloS one 2021-07, Vol.16 (7), p.e0254248
• Main Authors: Bearham, Jade, Krutrök, Nina, Lindberg, Botilda, Woodall, Maximillian, Astrand, Annika, Taylor, John D, Biggart, Matthew, Vasiljevs, Stanislavs, Tarran, Robert, Baines, Deborah L
• Format: Article
• Language: English
• Subjects: Analysis; Animals; Biology and Life Sciences; Biosensing Techniques - methods; Bronchus; Calcium-Binding Proteins; Chemical compounds; Dextrose; Diabetes; Diabetes mellitus; E coli; Engineering and Technology; Epithelial cells; Epithelium; Escherichia coli; Escherichia coli - metabolism; Ethylenediamine; Fluorescence; Fluorescent Dyes - chemistry; Fluorophores; Galactose; Genomics; Glucose; Glucose - metabolism; Hemodialysis; Humans; Immunology; Inhalation; Lungs; Measurement; Medicine and Health Sciences; Mice; Mice, Inbred C57BL; Molecular structure; Monosaccharide Transport Proteins - metabolism; Mutagenesis; Mutation; Periplasmic Binding Proteins - metabolism; Physical Sciences; Plasmids; Protein expression; Proteins; R&D; Research & development; Research and Analysis Methods; Respiration; Respiratory Mucosa - metabolism; Respiratory tract; Sensors; Staphylococcus infections
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0254248

We have modified the periplasmic Escherichia coli glucose/galactose binding protein (GBP) and labelled with environmentally sensitive fluorophores to further explore its potential as a sensor for the evaluation of glucose concentration in airway surface liquid (ASL). We identified E149C/A213R GBP labelled with N,N'-Dimethyl-N-(iodoacetyl)-N'-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)ethylenediamine (IANBD, emission wavelength maximum 536nm) with a Kd for D-glucose of 1.02mM and a fluorescence dynamic range of 5.8. This sensor was specific for D-glucose and exhibited fluorescence stability in experiments for several hours. The use of E149C/A213R GBP-IANBD in the ASL of airway cells grown at air-liquid-interface (ALI) detected an increase in glucose concentration 10 minutes after raising basolateral glucose from 5 to 15mM. This sensor also reported a greater change in ASL glucose concentration in response to increased basolateral glucose in H441 airway cells compared to human bronchial epithelial cells (HBEC) and there was less variability with HBEC data than that of H441 indicating that HBEC more effectively regulate glucose movement into the ASL. The sensor detected glucose in bronchoalveolar lavage fluid (BALf) from diabetic db/db mice but not normoglycaemic wildtype mice, indicating limited sensitivity of the sensor at glucose concentrations