Using a kinase-inducible bimolecular switch to control enzyme activity in living cells.

Sohum Mehta, Jin Zhang
Author Information
  1. Sohum Mehta: Department of Pharmacology and Molecular Sciences, The Johns Hopkins University School of Medicine, Baltimore, Maryland.

Abstract

Molecular switches have been instrumental in the development of powerful and versatile genetic tools for directly probing biochemical processes, such as intracellular signaling, within their native contexts. A molecular switch can be broadly defined as a molecular system capable of existing in either of two states (e.g., conformations), which can be converted from one state to the other by a specific input stimulus. This protocol outlines a method for using a kinase-inducible bimolecular switch, along with live-cell fluorescence microscopy, to directly control and monitor the activity of a specific enzyme in living cells.

Keywords

References

  1. ACS Chem Biol. 2012 Dec 21;7(12):1950-5 [PMID: 22999378]
  2. Structure. 2001 Mar 7;9(3):R33-8 [PMID: 11286893]
  3. Nature. 2009 Sep 3;461(7260):104-8 [PMID: 19693014]
  4. J Am Chem Soc. 2011 Apr 20;133(15):5676-9 [PMID: 21438554]
  5. PLoS One. 2008 Aug 27;3(8):e3068 [PMID: 18728784]
  6. ACS Chem Biol. 2013 Jan 18;8(1):116-21 [PMID: 23072367]
  7. Nat Methods. 2005 Jun;2(6):415-8 [PMID: 15908919]
  8. Sci Signal. 2011 Dec 13;4(203):ra87 [PMID: 22169478]
  9. Trends Biochem Sci. 1996 Nov;21(11):418-22 [PMID: 8987395]
  10. Annu Rev Biochem. 2011;80:375-401 [PMID: 21495849]
  11. Sci STKE. 2003 Jul 15;2003(191):RE12 [PMID: 12865499]
  12. Nat Methods. 2010 Mar;7(3):206-8 [PMID: 20154678]
  13. Chem Rev. 2011 May 11;111(5):3614-66 [PMID: 21456512]
  14. Curr Opin Cell Biol. 2001 Apr;13(2):131-8 [PMID: 11248545]
  15. Proc Natl Acad Sci U S A. 2012 Dec 26;109(52):E3668-77 [PMID: 23185021]

Grants

  1. DP1 CA174423/NCI NIH HHS
  2. R01 DK073368/NIDDK NIH HHS

MeSH Term

Cell Line
Cell Physiological Phenomena
DNA
Genes, Switch
Humans
Indicators and Reagents
Microscopy, Fluorescence
Phosphotransferases
Signal Transduction

Chemicals

Indicators and Reagents
DNA
Phosphotransferases

Word Cloud

Created with Highcharts 10.0.0switchdirectlybiochemicalmolecularcanspecifickinase-induciblebimolecularlive-cellfluorescencecontrolactivityenzymelivingcellsMolecularswitchesinstrumentaldevelopmentpowerfulversatilegenetictoolsprobingprocessesintracellularsignalingwithinnativecontextsbroadlydefinedsystemcapableexistingeithertwostatesegconformationsconvertedonestateinputstimulusprotocoloutlinesmethodusingalongmicroscopymonitorUsingperturbationbiosensorimaging

Similar Articles

Cited By

No available data.