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Real-time measurement of the intracellular pH of yeast cells during glucose metabolism using ratiometric fluorescent nanosensors

Elsutohy, Mohamed M.; Chauhan, Veeren M.; Markus, Robert; Kyyaly, Mohammed Aref; Tendler, Saul J.B.; Aylott, Jonathan W.

Authors

Mohamed M. Elsutohy

Veeren M. Chauhan

Robert Markus

Mohammed Aref Kyyaly

Saul J.B. Tendler

Jonathan W. Aylott



Abstract

Intracellular pH is a key parameter that influences many biochemical and metabolic pathways that can also be used as an indirect marker to monitor metabolic and intracellular processes. Herein, we utilise ratiometric fluorescent pH-sensitive nanosensors with an extended dynamic pH range to measure the intracellular pH of yeast (Saccharomyces cerevisiae) during glucose metabolism in real-time. Ratiometric fluorescent pH-sensitive nanosensors consisting of a polyacrylamide nanoparticle matrix covalently linked to two pH-sensitive fluorophores, Oregon green (OG) and 5(6)carboxyfluorescein (FAM), and a reference pH-insensitive fluorophore, 5(6)carboxytetramethylrhodamine (TAMRA), were synthesised. Nanosensors were functionalised with acrylamidopropyltrimethyl ammonium hydrochloride (ACTA) to confer a positive charge to the nanoparticle surfaces that facilitated nanosensor delivery to yeast cells, negating the need to use stress inducing techniques. The results showed that under glucose-starved conditions the intracellular pH of yeast population (n ≈ 200) was 4.67 ± 0.15. Upon addition of D-(+)-glucose (10 mM), this pH value decreased to pH 3.86 ± 0.13 over a period of 10 minutes followed by a gradual rise to a maximal pH of 5.21 ± 0.26, 25 minutes after glucose addition. 45 minutes after the addition of glucose, the intracellular pH of yeast cells returned to that of the glucose starved conditions. This study advances our understanding of the interplay between glucose metabolism and pH regulation in yeast cells, and indicates that the intracellular pH homestasis in yeast is highly regulated and demonstrates the utility of nanosensors for real-time intracellular pH measurements.

Journal Article Type Article
Publication Date May 14, 2017
Journal Nanoscale
Print ISSN 2040-3364
Electronic ISSN 2040-3372
Publisher Royal Society of Chemistry
Peer Reviewed Peer Reviewed
Volume 9
Issue 18
Pages 5904-5911
APA6 Citation Elsutohy, M. M., Chauhan, V. M., Markus, R., Kyyaly, M. A., Tendler, S. J., & Aylott, J. W. (2017). Real-time measurement of the intracellular pH of yeast cells during glucose metabolism using ratiometric fluorescent nanosensors. Nanoscale, 9(18), 5904-5911. https://doi.org/10.1039/c7nr00906b
DOI https://doi.org/10.1039/c7nr00906b
Publisher URL http://pubs.rsc.org/en/Content/ArticleLanding/2017/NR/C7NR00906B#!divAbstract
Copyright Statement Copyright information regarding this work can be found at the following address: http://eprints.nottingh.../end_user_agreement.pdf

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170531 Yeast Paper Accepted Manuscript.pdf (677 Kb)
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Copyright Statement
Copyright information regarding this work can be found at the following address: http://eprints.nottingham.ac.uk/end_user_agreement.pdf


170531 Yeast Paper Supporting Information.pdf (1.2 Mb)
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Copyright Statement
Copyright information regarding this work can be found at the following address: http://eprints.nottingham.ac.uk/end_user_agreement.pdf





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