Photo Control of Protein Function Using Photoactive Yellow Protein

  • Jakeb M. Reis
  • G. Andrew WoolleyEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1408)


Photoswitchable proteins are becoming increasingly common tools for manipulating cellular processes with high spatial and temporal precision. Photoactive yellow protein (PYP) is a small, water-soluble protein that undergoes a blue light induced change in conformation. It can serve as a scaffold for designing new tools to manipulate biological processes, but with respect to other protein scaffolds it presents some technical challenges. Here, we present practical information on how to overcome these, including how to synthesize the PYP chromophore, how to express and purify PYP, and how to screen for desired activity.

Key words

Photoactive yellow protein PYP Optogenetic Protein design Protein engineering 



This work has been supported by the Natural Sciences and Engineering Research Council of Canada and by NIH (R01MH086379).


  1. 1.
    Moglich A, Moffat K (2011) Engineered photoreceptors as novel optogenetic tools. Photochem Photobiol Sci 9(10):1286–1300CrossRefGoogle Scholar
  2. 2.
    Ramachandran PL et al (2011) The short-lived signaling state of the photoactive yellow protein photoreceptor revealed by combined structural probes. J Am Chem Soc 133(24):9395–9404CrossRefPubMedGoogle Scholar
  3. 3.
    Fan HY et al (2011) Improving a designed photocontrolled DNA-binding protein. Biochemistry 50(7):1226–1237CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Morgan SA, Woolley GA (2010) A photoswitchable DNA-binding protein based on a truncated GCN4-photoactive yellow protein chimera. Photochem Photobiol Sci 9(10):1320–1326CrossRefPubMedGoogle Scholar
  5. 5.
    Ali AM et al (2015) Optogenetic control of CREB signaling. Chem Biol. 22(11): 1531–9Google Scholar
  6. 6.
    Reis JM, Burns DC, Woolley GA (2014) Optical control of protein-protein interactions via blue light induced domain swapping. Biochemistry 53(30):5008–5016CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Kumar AB, Burns DC, Al-Abdul-Wahid MS, Woolley GA (2013) A circularly permuted photoactive yellow protein as a scaffold for photoswitch design. Biochemistry 52(19):3320–3331CrossRefPubMedGoogle Scholar
  8. 8.
    Changenet-Barret P et al (2002) Excited-state relaxation dynamics of a PYP chromophore model in solution: influence of the thioester group. Chem Phys Lett 365(3–4):285–291CrossRefGoogle Scholar
  9. 9.
    Genick UK et al (1997) Active site mutants implicate key residues for control of color and light cycle kinetics of photoactive yellow protein. Biochemistry 36(1):8–14CrossRefPubMedGoogle Scholar
  10. 10.
    Kyndt JA et al (2003) Heterologous production of Halorhodospira halophila holo-photoactive yellow protein through tandem expression of the postulated biosynthetic genes. Biochemistry 42(4):965–970CrossRefPubMedGoogle Scholar
  11. 11.
    Kumar A, Woolley GA (2015) Origins of the intermediate spectral form in M100 mutants of photoactive yellow protein. Photochem Photobiol 91(4):985–991CrossRefPubMedGoogle Scholar
  12. 12.
    Sasaki J, Kumauchi M, Hamada N, Oka T, Tokunaga F (2002) Light-induced unfolding of photoactive yellow protein mutant M100L. Biochemistry 41(6):1915–1922CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  1. 1.Department of ChemistryUniversity of TorontoTorontoCanada

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