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Cited by 475

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A genetically encoded fluorescent reporter of ATP:ADP ratio

Open access
Berg, Jim, Hung, Yin Pun, Yellen, Gary·2009·Nature Methods

Abstract

We constructed a fluorescent sensor of adenylate nucleotides by combining a circularly permuted variant of GFP with a bacterial regulatory protein, GlnK1, from Methanococcus jannaschii. The sensor's affinity for Mg-ATP was <100 nM, as seen for other members of the bacterial PII regulator family, a surprisingly high affinity given that normal intracellular ATP concentration is in the millimolar range. ADP bound the same site of the sensor as Mg-ATP, competing with it, but produced a smaller change in fluorescence. At physiological ATP and ADP concentrations, the binding site is saturated, but competition between the two substrates causes the sensor to behave as a nearly ideal reporter of the ATP:ADP concentration ratio. This principle for sensing the ratio of two analytes by competition at a high-affinity site probably underlies the normal functioning of PII regulatory proteins. The engineered sensor, Perceval, can be used to monitor the ATP:ADP ratio during live-cell imaging.

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Metadata

Authors
Berg, J., Hung, Y. P., & Yellen, G.
Venue
Nature Methods
Published
2009-01-04
Volume
6
Issue
2
Pages
161-166
ISSN
  • 1548-7091 (ISSN-L)
  • 1548-7105
OpenAlex
W2129558617
Cited by
475
References
1
FWCI
12.75
Type
journal article
Language
en
Open access
green

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