Optical Control of Cellular ATP Levels with a Photocaged Adenylate Kinase

We have developed a new tool for the optical control of cellular ATP concentrations with a photocaged adenylate kinase (Adk). The photocaged Adk is generated by substituting a catalytically essential lysine with a hydroxycoumarin‐protected lysine through site‐specific unnatural amino acid mutagenesi...

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Bibliographic Details
Published in:Chembiochem : a European journal of chemical biology 2020-07, Vol.21 (13), p.1832-1836
Main Authors: Zhou, Wenyuan, Hankinson, Chasity P., Deiters, Alexander
Format: Article
Language:English
Subjects:
Adenylate kinase
Amino acids
ATP
ATP homeostasis
caged compounds
Depletion
E coli
Kinases
Lysine
Mammalian cells
Mammals
Mutagenesis
Optical control
Perturbation
Phosphotransferase
photocaging
Restoration
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Summary:We have developed a new tool for the optical control of cellular ATP concentrations with a photocaged adenylate kinase (Adk). The photocaged Adk is generated by substituting a catalytically essential lysine with a hydroxycoumarin‐protected lysine through site‐specific unnatural amino acid mutagenesis in both E. coli and mammalian cells. Caging of the critical lysine residue offers complete suppression of Adk's phosphotransferase activity and rapid restoration of its function both in vitro and in vivo upon optical stimulation. Light‐activated Adk renders faster rescue of cell growth than chemically inducible expression of wild‐type Adk in E. coli as well as rapid ATP depletion in mammalian cells. Thus, caging Adk provides a new tool for direct conditional perturbation of cellular ATP concentrations thereby enabling the investigation of ATP‐coupled physiological events in temporally dynamic contexts. Direct optical perturbation of cellular ATP levels is achieved by substituting the essential lysine of E. coli adenylate kinase with a hydroxycoumarin‐protected lysine through unnatural amino acid mutagenesis. The caged lysine residue renders the kinase inactive until photolysis reactivates it in both E. coli and mammalian cells.
ISSN:1439-4227
1439-7633
DOI:10.1002/cbic.201900757