Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Review Article
  • Published:

Using azobenzene photocontrol to set proteins in motion

Nature Reviews Chemistry volume 6, pages 112–124 (2022)Cite this article

Subjects

Abstract

Controlling the activity of proteins with azobenzene photoswitches is a potent tool for manipulating their biological function. With the help of light, it is possible to change binding affinities, control allostery or manipulate complex biological processes, for example. Additionally, owing to their intrinsically fast photoisomerization, azobenzene photoswitches can serve as triggers that initiate out-of-equilibrium processes. Such switching of the activity initiates a cascade of conformational events that can be accessed with time-resolved methods. In this Review, we show how the potency of azobenzene photoswitching can be combined with transient spectroscopic techniques to disclose the order of events and experimentally observe biomolecular interactions in real time. This strategy will further our understanding of how a protein can accommodate, adapt and readjust its structure to answer an incoming signal, revealing more of the dynamical character of proteins.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: Strategies of azobenzene photocontrol.
Fig. 2: Artificially photoswitchable proteins studied in the context of protein allostery and protein–ligand binding by time-resolved spectroscopy.
Fig. 3: Typical example of a time-resolved data set and its timescale analysis.
Fig. 4: Binding curves for different photoswitchable S-peptide variants to S-protein.
Fig. 5: Typical timescales.
Fig. 6: Photoswitchable ATPase.

Similar content being viewed by others

References

  1. Henzler-Wildman, K. & Kern, D. Dynamic personalities of proteins. Nature 450, 964–972 (2007).

    Article  CAS  PubMed  Google Scholar 

  2. Dyson, H. J. & Wright, P. E. Equilibrium NMR studies of unfolded and partially folded proteins. Nat. Struct. Biol. 5, 499–503 (1998).

    Article  CAS  PubMed  Google Scholar 

  3. Bernado, P. & Blackledge, M. Proteins in dynamic equilibrium. Nature 468, 1046–1048 (2010).

    Article  CAS  PubMed  Google Scholar 

  4. Elber, R. & Karplus, M. Enhanced sampling in molecular dynamics: use of the time-dependent Hartree approximation for a simulation of carbon monoxide diffusion through myoglobin. J. Am. Chem. Soc. 112, 9161–9175 (1990).

    Article  CAS  Google Scholar 

  5. Palmer, A. G. NMR characterization of the dynamics of biomacromolecules. Chem. Rev. 104, 3623–3640 (2004).

    Article  CAS  PubMed  Google Scholar 

  6. Mittermaier, A. & Kay, L. E. New tools provide new insights in NMR studies of protein dynamics. Science 312, 224–228 (2006).

    Article  CAS  PubMed  Google Scholar 

  7. Kay, L. E. NMR studies of protein structure and dynamics. J. Magn. Reson. 173, 193–207 (2005).

    Article  CAS  PubMed  Google Scholar 

  8. Olsson, M. H., Parson, W. W. & Warshel, A. Dynamical contributions to enzyme catalysis: critical tests of a popular hypothesis. Chem. Rev. 106, 1737–1756 (2006).

    Article  CAS  PubMed  Google Scholar 

  9. Benkovic, S. J. & Hammes-Schiffer, S. A perspective on enzyme catalysis. Science 301, 1196–1202 (2003).

    Article  CAS  PubMed  Google Scholar 

  10. Csermely, P., Palotai, R. & Nussinov, R. Induced fit, conformational selection and independent dynamic segments: an extended view of binding events. Trends Biochem. Sci. 35, 539–546 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Swain, J. F. & Gierasch, L. M. The changing landscape of protein allostery. Curr. Opin. Struct. Biol. 16, 102–108 (2006).

    Article  CAS  PubMed  Google Scholar 

  12. Tsai, C. J., del Sol, A. & Nussinov, R. Allostery: absence of a change in shape does not imply that allostery is not at play. J. Mol. Biol. 378, 1–11 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Smock, R. G. & Gierasch, L. M. Sending signals dynamically. Science 324, 198–203 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Tsai, C. J. & Nussinov, R. A unified view of “how allostery works”. PLoS Comput. Biol. 10, e1003394 (2014).

    Article  PubMed  PubMed Central  Google Scholar 

  15. Hilser, V. J., Wrabl, J. O. & Motlagh, H. N. Structural and energetic basis of allostery. Annu. Rev. Biophys. 41, 585–609 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Schotte, F. et al. Watching a protein as it functions with 150-ps time-resolved X-ray crystallography. Science 300, 1944–1947 (2003).

    Article  CAS  PubMed  Google Scholar 

  17. Knapp, J. E., Pahl, R., Šrajer, V. & Royer, W. E. Allosteric action in real time: time-resolved crystallographic studies of a cooperative dimeric hemoglobin. Proc. Natl Acad. Sci. USA 103, 7649–7654 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Kern, J. et al. Simultaneous femtosecond X-ray spectroscopy and diffraction of photosystem II at room temperature. Science 340, 491–496 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Nogly, P. et al. Retinal isomerization in bacteriorhodopsin captured by a femtosecond X-ray laser. Science 361, eaat0094 (2018).

    Article  PubMed  Google Scholar 

  20. Standfuss, J. Membrane protein dynamics studied by X-ray lasers — or why only time will tell. Curr. Opin. Struct. Biol. 57, 63–71 (2019).

    Article  CAS  PubMed  Google Scholar 

  21. Skopintsev, P. et al. Femtosecond-to-millisecond structural changes in a light-driven sodium pump. Nature 583, 314–318 (2020).

    Article  CAS  PubMed  Google Scholar 

  22. Chapman, H. N. et al. Femtosecond X-ray protein nanocrystallography. Nature 470, 73–77 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Wang, Q., Schoenlein, R. W., Peteanu, L. A., Mathies, R. A. & Shank, C. V. Vibrationally coherent photochemistry in the femtosecond primary event of vision. Science 266, 422–424 (1994).

    Article  CAS  PubMed  Google Scholar 

  24. Zinth, W. & Wachtveitl, J. The first picoseconds in bacterial photosynthesis-ultrafast electron transfer for the efficient conversion of light energy. ChemPhysChem 6, 871–880 (2005).

    Article  CAS  PubMed  Google Scholar 

  25. Palczewski, K. G protein-coupled receptor rhodopsin. Annu. Rev. Biochem. 75, 743–767 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Engel, G. S. et al. Evidence for wavelike energy transfer through quantum coherence in photosynthetic systems. Nature 446, 782–786 (2007).

    Article  CAS  PubMed  Google Scholar 

  27. Mirkovic, T. et al. Light absorption and energy transfer in the antenna complexes of photosynthetic organisms. Chem. Rev. 117, 249–293 (2017).

    Article  CAS  PubMed  Google Scholar 

  28. Hegemann, P. Algal sensory photoreceptors. Annu. Rev. Plant. Biol. 59, 167–189 (2008).

    Article  CAS  PubMed  Google Scholar 

  29. Rockwell, N. C. & Lagarias, J. C. Phytochrome diversification in cyanobacteria and eukaryotic algae. Curr. Opin. Plant. Biol. 37, 87–93 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Austin, R. H., Beeson, K. W., Eisenstein, L., Frauenfelder, H. & Gunsalus, I. C. Dynamics of ligand binding to myoglobin. Biochemistry 14, 5355–5373 (1975).

    Article  CAS  PubMed  Google Scholar 

  31. Cammarata, M., Levantino, M., Wulff, M. & Cupane, A. Unveiling the timescale of the R-T transition in human hemoglobin. J. Mol. Biol. 400, 951–962 (2010).

    Article  CAS  PubMed  Google Scholar 

  32. Ellis-Davies, G. C. Caged compounds: photorelease technology for control of cellular chemistry and physiology. Nat. Methods 4, 619–628 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Causgrove, T. P. & Dyer, R. B. Nonequilibrium protein folding dynamics: laser induced pH-jump studies of the helix–coil transition. Chem. Phys. 323, 2–10 (2006).

    Article  CAS  Google Scholar 

  34. Donten, M. L. et al. pH-Jump induced leucine zipper folding beyond the diffusion limit. J. Phys. Chem. B 119, 1425–1432 (2015).

    Article  CAS  PubMed  Google Scholar 

  35. Thompson, P. A., Eaton, W. A. & Hofrichter, J. Laser temperature jump study of the helix–coil kinetics of an alanine peptide interpreted with a ‘kinetic zipper’ model. Biochemistry 36, 9200–9210 (1997).

    Article  CAS  PubMed  Google Scholar 

  36. Callender, R. H., Dyer, R. B., Gilmanshin, R. & Woodruff, W. H. Fast events in protein folding: the time evolution of primary processes. Annu. Rev. Phys. Chem. 49, 173–202 (1998).

    Article  CAS  PubMed  Google Scholar 

  37. Yang, W. Y. & Gruebele, M. Folding at the speed limit. Nature 423, 193–197 (2003).

    Article  CAS  PubMed  Google Scholar 

  38. Hauser, K., Krejtschi, C., Huang, R., Wu, L. & Keiderling, T. A. Site-specific relaxation kinetics of a tryptophan zipper hairpin peptide using temperature-jump IR spectroscopy and isotopic labeling. J. Am. Chem. Soc. 130, 2984–2992 (2008).

    Article  CAS  PubMed  Google Scholar 

  39. Crespi, S., Simeth, N. A. & König, B. Heteroaryl azo dyes as molecular photoswitches. Nat. Rev. Chem. 3, 133–146 (2019).

    Article  CAS  Google Scholar 

  40. Blanco-Lomas, M., Samanta, S., Campos, P. J., Woolley, G. A. & Sampedro, D. Reversible photocontrol of peptide conformation with a rhodopsin-like photoswitch. J. Am. Chem. Soc. 134, 6960–6963 (2012).

    Article  CAS  PubMed  Google Scholar 

  41. Szymański, W. et al. Reversible photocontrol of biological systems by the incorporation of molecular photoswitches. Chem. Rev. 113, 6114–6178 (2013).

    Article  Google Scholar 

  42. Willner, I., Rubin, S. & Riklin, A. Photoregulation of papain activity through anchoring photochromic azo groups to the enzyme backbone. J. Am. Chem. Soc. 113, 3321–3325 (1991).

    Article  CAS  Google Scholar 

  43. James, D., Burns, D. C. & Woolley, G. Kinetic characterization of ribonuclease S mutants containing photoisomerizable phenylazophenylalanine residues. Protein Eng. Des. Sel. 14, 983–991 (2001).

    Article  CAS  Google Scholar 

  44. Liu, D., Karanicolas, J., Yu, C., Zhang, Z. & Woolley, G. A. Site-specific incorporation of photoisomerizable azobenzene groups into ribonuclease S. Bioorg. Med. Chem. Lett. 7, 2677–2680 (1997).

    Article  CAS  Google Scholar 

  45. Hamachi, I., Hiraoka, T., Yamada, Y. & Shinkai, S. Photoswitching of the enzymatic activity of semisynthetic ribonuclease S′ bearing phenylazophenylalanine at a specific site. Chem. Lett. 27, 537–538 (1998).

    Article  Google Scholar 

  46. Yamada, M. D., Nakajima, Y., Maeda, H. & Maruta, S. Photocontrol of kinesin ATPase activity using an azobenzene derivative. J. Biochem. 142, 691–698 (2007).

    Article  CAS  PubMed  Google Scholar 

  47. Schierling, B. et al. Controlling the enzymatic activity of a restriction enzyme by light. Proc. Natl Acad. Sci. USA 107, 1361–1366 (2010).

    Article  CAS  PubMed  Google Scholar 

  48. Ritterson, R. S., Kuchenbecker, K. M., Michalik, M. & Kortemme, T. Design of a photoswitchable cadherin. J. Am. Chem. Soc. 135, 12516–12519 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Buchli, B. et al. Kinetic response of a photoperturbed allosteric protein. Proc. Natl Acad. Sci. USA 110, 11725–11730 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Borowiak, M. et al. Photoswitchable inhibitors of microtubule dynamics optically control mitosis and cell death. Cell 162, 402–411 (2015).

    Article  Google Scholar 

  51. Hoersch, D. Engineering a light-controlled F 1 ATPase using structure-based protein design. PeerJ 4, e2286 (2016).

    Article  PubMed  PubMed Central  Google Scholar 

  52. Blacklock, K. M., Yachnin, B. J., Woolley, G. A. & Khare, S. D. Computational design of a photocontrolled cytosine deaminase. J. Am. Chem. Soc. 140, 14–17 (2018).

    Article  CAS  PubMed  Google Scholar 

  53. Bozovic, O., Jankovic, B. & Hamm, P. Sensing the allosteric force. Nat. Commun. 11, 5841 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  54. Bozovic, O. et al. The speed of allosteric signaling within a single-domain protein. J. Phys. Chem. Lett. 12, 4262–4267 (2021).

    Article  CAS  PubMed  Google Scholar 

  55. Jankovic, B. et al. Sequence of events during peptide unbinding from rnase s: a complete experimental description. J. Phys. Chem. Lett. 12, 5201–5207 (2021).

    Article  CAS  PubMed  Google Scholar 

  56. Guerrero, L. et al. Photochemical regulation of DNA-binding specificity of MyoD. Angew. Chem. Int. Ed. 44, 7778–7782 (2005).

    Article  CAS  Google Scholar 

  57. Guerrero, L., Smart, O. S., Woolley, G. A. & Allemann, R. K. Photocontrol of DNA binding specificity of a miniature engrailed homeodomain. J. Am. Chem. Soc. 127, 15624–15629 (2005).

    Article  CAS  PubMed  Google Scholar 

  58. Woolley, G. A. et al. Reversible photocontrol of DNA binding by a designed GCN4-bZIP protein. Biochemistry 45, 6075–6084 (2006).

    Article  CAS  PubMed  Google Scholar 

  59. Kneissl, S., Loveridge, E. J., Williams, C., Crump, M. P. & Allemann, R. K. Photocontrollable peptide-based switches target the anti-apoptotic protein Bcl-x L. ChemBioChem 9, 3046–3054 (2008).

    Article  CAS  PubMed  Google Scholar 

  60. Zhang, F., Timm, K. A., Arndt, K. M. & Woolley, G. A. Photocontrol of coiled-coil proteins in living cells. Angew. Chem. Int. Ed. 49, 3943–3946 (2010).

    Article  CAS  Google Scholar 

  61. Nevola, L. et al. Light-regulated stapled peptides to inhibit protein-protein interactions involved in clathrin-mediated endocytosis. Angew. Chem. Int. Ed. 52, 7704–7708 (2013).

    Article  CAS  Google Scholar 

  62. Martín-Quirós, A. et al. Absence of a stable secondary structure is not a limitation for photoswitchable inhibitors of β-arrestin/β-adaptin 2 protein–protein interaction. Chem. Biol. 22, 31–37 (2015).

    Article  PubMed  Google Scholar 

  63. Babalhavaeji, A. & Woolley, G. A. Modular design of optically controlled protein affinity reagents. Chem. Commun. 54, 1591–1594 (2018).

    Article  CAS  Google Scholar 

  64. Albert, L. et al. Modulating protein–protein interactions with visible-light-responsive peptide backbone photoswitches. ChemBioChem 20, 1417–1429 (2019).

    Article  CAS  PubMed  Google Scholar 

  65. Jankovic, B. et al. Photocontrolling protein–peptide interactions: from minimal perturbation to complete unbinding. J. Am. Chem. Soc. 141, 10702–10710 (2019).

    Article  CAS  PubMed  Google Scholar 

  66. Myrhammar, A., Rosik, D. & Karlström, A. E. Photocontrolled reversible binding between the protein A-derived Z domain and immunoglobulin G. Bioconjug. Chem. 31, 622–630 (2020).

    Article  CAS  PubMed  Google Scholar 

  67. Bozovic, O. et al. Real-time observation of ligand-induced allosteric transitions in a PDZ domain. Proc. Natl Acad. Sci. USA 117, 26031–26039 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  68. Day, K. et al. Photoinduced reconfiguration to control the protein-binding affinity of azobenzene-cyclized peptides. J. Mater. Chem. B 8, 7413–7427 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  69. Jankovic, B., Bozovic, O. & Hamm, P. Intrinsic dynamics of protein–peptide unbinding. Biochemistry 60, 1755–1763 (2021).

    Article  CAS  PubMed  Google Scholar 

  70. Beharry, A. A. & Woolley, G. A. Azobenzene photoswitches for biomolecules. Chem. Soc. Rev. 40, 4422–4437 (2011).

    Article  CAS  PubMed  Google Scholar 

  71. Borisenko, V. & Woolley, G. A. Reversibility of conformational switching in light-sensitive peptides. J. Photochem. Photobiol. A 173, 21–28 (2005).

    Article  CAS  Google Scholar 

  72. Nägele, T., Hoche, R., Zinth, W. & Wachtveitl, J. Femtosecond photoisomerization of cis-azobenzene. Chem. Phys. Lett. 272, 489–495 (1997).

    Article  Google Scholar 

  73. Dong, M., Babalhavaeji, A., Samanta, S., Beharry, A. A. & Woolley, G. A. Red-shifting azobenzene photoswitches for in vivo use. Acc. Chem. Res. 48, 2662–2670 (2015).

    Article  CAS  PubMed  Google Scholar 

  74. Dong, M. et al. Near-infrared photoswitching of azobenzenes under physiological conditions. J. Am. Chem. Soc. 139, 13483–13486 (2017).

    Article  CAS  PubMed  Google Scholar 

  75. Zhang, Z., Burns, D. C., Kumita, J. R., Smart, O. S. & Woolley, G. A. A water-soluble azobenzene cross-linker for photocontrol of peptide conformation. Bioconjugate Chem. 14, 824–829 (2003).

    Article  CAS  Google Scholar 

  76. Sadovski, O., Beharry, A. A., Zhang, F. & Woolley, G. A. Spectral tuning of azobenzene photoswitches for biological applications. Angew. Chem. Int. Ed. 48, 1484–1486 (2009).

    Article  CAS  Google Scholar 

  77. Broichhagen, J., Frank, J. A. & Trauner, D. A roadmap to success in photopharmacology. Acc. Chem. Res. 48, 1947–1960 (2015).

    Article  CAS  PubMed  Google Scholar 

  78. Ankenbruck, N., Courtney, T., Naro, Y. & Deiters, A. Optochemical control of biological processes in cells and animals. Angew. Chem. Int. Ed. 57, 2768–2798 (2018).

    Article  CAS  Google Scholar 

  79. Zhu, M. & Zhou, H. Azobenzene-based small molecular photoswitches for protein modulation. Org. Biomol. Chem. 16, 8434–8445 (2018).

    Article  CAS  PubMed  Google Scholar 

  80. Hüll, K., Morstein, J. & Trauner, D. In vivo photopharmacology. Chem. Rev. 118, 10710–10747 (2018).

    Article  PubMed  Google Scholar 

  81. Albert, L. & Vázquez, O. Photoswitchable peptides for spatiotemporal control of biological functions. Chem. Commun. 55, 10192–10213 (2019).

    Article  CAS  Google Scholar 

  82. Paoletti, P., Ellis-Davies, G. C. & Mourot, A. Optical control of neuronal ion channels and receptors. Nat. Rev. Neurosci. 20, 514–532 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  83. Peddie, V. & Abell, A. D. Photocontrol of peptide secondary structure through non-azobenzene photoswitches. J. Photochem. Photobiol. C 40, 1–20 (2019).

    Article  CAS  Google Scholar 

  84. Beharry, A. A., Wong, L., Tropepe, V. & Woolley, G. A. Fluorescence imaging of azobenzene photoswitching in vivo. Angew. Chem. Int. Ed. 50, 1325–1327 (2011).

    Article  CAS  Google Scholar 

  85. Behrendt, R. et al. Photomodulation of the conformation of cyclic peptides with azobenzene moieties in the peptide backbone. Angew. Chem. Int. Ed. 38, 2771–2774 (1999).

    Article  CAS  Google Scholar 

  86. Spörlein, S. et al. Ultrafast spectroscopy reveals subnanosecond peptide conformational dynamics and validates molecular dynamics simulation. Proc. Natl Acad. Sci. USA 99, 7998–8002 (2002).

    Article  PubMed  PubMed Central  Google Scholar 

  87. Bredenbeck, J. et al. Picosecond conformational transition and equilibration of a cyclic peptide. Proc. Natl Acad. Sci. USA 100, 6452–6457 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  88. Aemissegger, A., Krautler, V., van Gunsteren, W. F. & Hilvert, D. A photoinducible beta-hairpin. J. Am. Chem. Soc. 127, 2929–2936 (2005).

    Article  CAS  PubMed  Google Scholar 

  89. Rehm, S., Lenz, M. O., Mensch, S., Schwalbe, H. & Wachtveitl, J. Ultrafast spectroscopy of a photoswitchable 30-amino acid de novo synthesized peptide. Chem. Phys. 323, 28–35 (2005).

    Article  Google Scholar 

  90. Schrader, T. E. et al. Light-triggered β-hairpin folding and unfolding. Proc. Natl Acad. Sci. USA 104, 15729–15734 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  91. Rampp, M. S. et al. Time-resolved infrared studies of the unfolding of a light triggered β-hairpin peptide. Chem. Phys. 512, 116–121 (2018).

    Article  CAS  Google Scholar 

  92. Kumita, J. R., Smart, O. S. & Woolley, G. A. Photo-control of helix content in a short peptide. Proc. Natl Acad. Sci. USA 97, 3803–3808 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  93. Flint, D. G., Kumita, J. R., Smart, O. S. & Woolley, G. A. Using an azobenzene cross-linker to either increase or decrease peptide helix content upon trans-to-cis photoisomerization. Chem. Biol. 9, 391–397 (2002).

    Article  CAS  PubMed  Google Scholar 

  94. Woolley, G. A. Photocontrolling peptide alpha helices. Acc. Chem. Res. 38, 486–493 (2005).

    Article  CAS  PubMed  Google Scholar 

  95. Chen, E., Kumita, J. R., Woolley, G. A. & Kliger, D. S. The kinetics of helix unfolding of an azobenzene cross-linked peptide probed by nanosecond time-resolved optical rotatory dispersion. J. Am. Chem. Soc. 125, 12443–12449 (2003).

    Article  CAS  PubMed  Google Scholar 

  96. Bredenbeck, J., Helbing, J., Kumita, J. R., Woolley, G. A. & Hamm, P. α-Helix formation in a photoswitchable peptide tracked from picoseconds to microseconds by time resolved IR spectroscopy. Proc. Natl Acad. Sci. USA 102, 2379–2384 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  97. Ihalainen, J. A. et al. Folding and unfolding of a photoswitchable peptide from picoseconds to microseconds. Proc. Natl Acad. Sci. USA 104, 5383–5388 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  98. Ihalainen, J. A. et al. α-Helix folding in the presence of structural constraints. Proc. Natl Acad. Sci. USA 105, 9588–9593 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  99. Zhang, F. et al. Structure-based approach to the photocontrol of protein folding. J. Am. Chem. Soc. 131, 2283–2289 (2009).

    Article  CAS  PubMed  Google Scholar 

  100. Waldauer, S. A., Stucki-Buchli, B., Frey, L. & Hamm, P. Effect of viscogens on the kinetic response of a photoperturbed allosteric protein. J. Chem. Phys. 141, 22D514 (2014).

    Article  PubMed  Google Scholar 

  101. Lee, H.-J. & Zheng, J. J. PDZ domains and their binding partners: structure, specificity, and modification. Cell Commun. Signal. 8, 8 (2010).

    Article  PubMed  PubMed Central  Google Scholar 

  102. Ivarsson, Y. Plasticity of PDZ domains in ligand recognition and signaling. FEBS Lett. 586, 2638–2647 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  103. Morais Cabral, J. H. et al. Crystal structure of a PDZ domain. Nature 382, 649–652 (1996).

    Article  CAS  PubMed  Google Scholar 

  104. Basdevant, N., Weinstein, H. & Ceruso, M. Thermodynamic basis for promiscuity and selectivity in protein-protein interactions: PDZ domains, a case study. J. Am. Chem. Soc. 128, 12766–12777 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  105. Münz, M., Hein, J. & Biggin, P. C. The role of flexibility and conformational selection in the binding promiscuity of PDZ domains. PLoS Comput. Biol. 8, e1002749 (2012).

    Article  PubMed  PubMed Central  Google Scholar 

  106. Gerek, Z. N., Keskin, O. & Ozkan, S. B. Identification of specificity and promiscuity of PDZ domain interactions through their dynamic behavior. Proteins 77, 796–811 (2009).

    Article  CAS  PubMed  Google Scholar 

  107. Tonikian, R. et al. A specificity map for the PDZ domain family. PLoS Biol. 6, 2043–2059 (2008).

    Article  CAS  Google Scholar 

  108. te Velthuis, A. J., Sakalis, P. A., Fowler, D. A. & Bagowski, C. P. Genome-wide analysis of PDZ domain binding reveals inherent functional overlap within the PDZ interaction network. PLoS ONE 6, e16047 (2011).

    Article  Google Scholar 

  109. Petit, C. M., Zhang, J., Sapienza, P. J., Fuentes, E. J. & Lee, A. L. Hidden dynamic allostery in a PDZ domain. Proc. Natl Acad. Sci. USA 106, 18249–18254 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  110. Fuentes, E. J., Gilmore, S. A., Mauldin, R. V. & Lee, A. L. Evaluation of energetic and dynamic coupling networks in a PDZ domain protein. J. Mol. Biol. 364, 337–351 (2006).

    Article  CAS  PubMed  Google Scholar 

  111. Li, J., Callaway, D. J. E. & Bu, Z. Ezrin induces long-range interdomain allostery in the scaffolding protein NHERF1. J. Mol. Biol. 392, 166–180 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  112. Fuentes, E. J., Der, C. J. & Lee, A. L. Ligand-dependent dynamics and intramolecular signaling in a PDZ domain. J. Mol. Biol. 335, 1105–1115 (2004).

    Article  CAS  PubMed  Google Scholar 

  113. Richards, F. M. On the enzymic activity of subtilisin-modified ribonuclease. Proc. Natl Acad. Sci. USA 44, 162–166 (1958).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  114. Roberts, G. C. K., Dennis, E. A., Mwadow, D. H., Cohen, J. S. & Oleg, J. The mechanism of action of ribonuclease. Proc. Natl Acad. Sci. USA 62, 1151–1158 (1969).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  115. Richards, F. M. & Vithayathil, P. J. The preparation of subtilisin-modified ribonuclease and the separation of the peptide and protein components. J. Biol. Chem. 234, 1459–1465 (1959).

    Article  CAS  PubMed  Google Scholar 

  116. Wlodawer, A. & Sjolin, L. Hydrogen exchange in RNase A: neutron diffraction study. Proc. Natl Acad. Sci. USA 79, 1418–1422 (1982).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  117. Goldberg, J. M. & Baldwin, R. L. A specific transition state for S-peptide combining with folded S-protein and then refolding. Proc. Natl Acad. Sci. USA 96, 2019–2024 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  118. Bachmann, A., Wildemann, D., Praetorius, F., Fischer, G. & Kiefhaber, T. Mapping backbone and side-chain interactions in the transition state of a coupled protein folding and binding reaction. Proc. Natl Acad. Sci. USA 108, 3952–3957 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  119. Luitz, M. P., Bomblies, R. & Zacharias, M. Comparative molecular dynamics analysis of RNase-S complex formation. Biophys. J. 113, 1466–1474 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  120. Schreier, A. A. & Baldwin, R. L. Mechanism of dissociation of s-peptide from ribonuclease. Biochemistry 16, 4203–4209 (1977).

    Article  CAS  PubMed  Google Scholar 

  121. Hamm, P., Ohline, S. M. & Zinth, W. Vibrational cooling after ultrafast photoisomerization of azobenzene measured by femtosecond infrared spectroscopy. J. Chem. Phys. 106, 519–529 (1997).

    Article  CAS  Google Scholar 

  122. Baumann, T. et al. Site-resolved observation of vibrational energy transfer using a genetically encoded ultrafast heater. Angew. Chem. Int. Ed. 58, 2899–2903 (2019).

    Article  CAS  Google Scholar 

  123. Barth, A. & Zscherp, C. What vibrations tell us about proteins. Quart. Rev. Biophys. 35, 369–430 (2002).

    Article  CAS  Google Scholar 

  124. Frauenfelder, H., Sligar, S. G. & Wolynes, P. G. The energy landscapes and motions of proteins. Science 254, 1598–1603 (1991).

    Article  CAS  PubMed  Google Scholar 

  125. Dill, K. A. & Chan, H. S. From levinthal to pathways to funnels: the “new view” of protein folding kinetics. Nat. Struct. Biol. 4, 10–19 (1997).

    Article  CAS  PubMed  Google Scholar 

  126. Stock, G. & Hamm, P. A nonequilibrium approach to allosteric communication. Phil. Trans. R. Soc. B 373, 20170187 (2018).

    Article  PubMed  PubMed Central  Google Scholar 

  127. Buchenberg, S., Sittel, F. & Stock, G. Time-resolved observation of protein allosteric communication. Proc. Natl Acad. Sci. USA 114, E6804–E6811 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  128. Buhrke, D., Oppelt, K. T., Heckmeier, P. J., Fernandez-Teran, R. & Hamm, P. Nanosecond protein dynamics in a red/green cyanobacteriochrome revealed by transient IR spectroscopy. J. Chem. Phys. 153, 245101 (2020).

    Article  CAS  PubMed  Google Scholar 

  129. Doyle, D. A. et al. Crystal structures of a complexed and peptide-free membrane protein- binding domain: molecular basis of peptide recognition by PDZ. Cell 85, 1067–1076 (1996).

    Article  CAS  PubMed  Google Scholar 

  130. Ballif, B. A., Carey, G. R., Sunyaev, S. R. & Gygi, S. P. Large-scale identification and evolution indexing of tyrosine phosphorylation sites from murine brain. J. Proteome Res. 7, 311–318 (2008).

    Article  CAS  PubMed  Google Scholar 

  131. Zhang, J., Petit, C. M., King, D. S. & Lee, A. L. Phosphorylation of a PDZ domain extension modulates binding affinity and interdomain interactions in postsynaptic density-95 (PSD-95) protein, a membrane-associated guanylate kinase (MAGUK). J. Biol. Chem. 286, 41776–41785 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  132. Boehr, D. D., Nussinov, R. & Wright, P. E. The role of dynamic conformational ensembles in biomolecular recognition. Nat. Chem. Biol. 5, 789–796 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  133. Morando, M. A. et al. Conformational selection and induced fit mechanisms in the binding of an anticancer drug to the c-Src kinase. Sci. Rep. 6, 24439 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  134. Vogt, A. D. & DiCera, E. Conformational selection or induced fit? A critical appraisal of the kinetic mechanism. Biochemistry 41, 5894–5902 (2012).

    Article  Google Scholar 

  135. Gianni, S., Dogan, J. & Jemth, P. Distinguishing induced fit from conformational selection. Biophys. Chem. 189, 33–39 (2014).

    Article  CAS  PubMed  Google Scholar 

  136. Hammes, G. G., Chang, Y.-C. & Oas, T. G. Conformational selection or induced fit: a flux description of reaction mechanism. Proc. Natl Acad. Sci. USA 106, 13737–13741 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  137. Deisseroth, K. Optogenetics. Nat. Methods 8, 26–29 (2011).

    Article  CAS  PubMed  Google Scholar 

  138. Häusser, M. Optogenetics: the age of light. Nat. Methods 11, 1012–1014 (2014).

    Article  PubMed  Google Scholar 

  139. Hoppmann, C. et al. Genetically encoding photoswitchable click amino acids in Escherichia coli and mammalian cells. Angew. Chem. Int. Ed. 53, 3932–3936 (2014).

    Article  CAS  Google Scholar 

  140. Hoppmann, C., Maslennikov, I., Choe, S. & Wang, L. In situ formation of an azo bridge on proteins controllable by visible light. J. Am. Chem. Soc. 137, 11218–11221 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  141. Koziol, K. L., Johnson, P. J. M., Stucki-Buchli, B., Waldauer, S. A. & Hamm, P. Fast infrared spectroscopy of protein dynamics: advancing sensitivity and selectivity. Curr. Opin. Struct. Biol. 34, 1–6 (2015).

    Article  CAS  PubMed  Google Scholar 

  142. Murphy, R. E., Cook, F. H. & Sakai, H. Time-resolved Fourier spectroscopy. J. Opt. Soc. Am. 65, 600–604 (1975).

    Article  CAS  Google Scholar 

  143. Uhmann, W., Becker, A., Taran, C. & Siebert, F. Time-resolved FT-IR absorption-spectroscopy using a step-scan interferometer. Appl. Spectrosc. 45, 390–397 (1991).

    Article  CAS  Google Scholar 

  144. Gerwert, K. Molecular reaction mechanisms of proteins monitored by time-resolved FT-IR difference. Biol. Chem. 380, 931–935 (1999).

    Article  CAS  PubMed  Google Scholar 

  145. Radu, I., Schleeger, M., Bolwien, C. & Heberle, J. Time-resolved methods in biophysics. 10. Time-resolved FT-IR difference spectroscopy and the application to membrane proteins. Photochem. Photobiol. Sci. 8, 1517–1528 (2009).

    Article  CAS  PubMed  Google Scholar 

  146. Ritter, E. et al. Time-resolved infrared spectroscopic techniques as applied to channel rhodopsin. Front. Mol. Biosci. 2, 38 (2015).

    Article  PubMed  PubMed Central  Google Scholar 

  147. Zhang, L., Tian, G., Li, J. & Yu, B. Applications of absorption spectroscopy using quantum cascade lasers. Appl. Spectrosc. 68, 1095–1107 (2014).

    Article  CAS  PubMed  Google Scholar 

  148. Schultz, B. J., Mohrmann, H., Lorenz-Fonfria, V. A. & Heberle, J. Protein dynamics observed by tunable mid-IR quantum cascade lasers across the time range from 10 ns to 1 s. Spectrochim. Acta A 188, 666–674 (2018).

    Article  CAS  Google Scholar 

  149. Stritt, P., Jawurek, M. & Hauser, K. Application of tunable quantum cascade lasers to monitor dynamics of bacteriorhodopsin in the mid-IR spectral range. Biomed. Spectrosc. Imaging 9, 55–61 (2020).

    Article  CAS  Google Scholar 

  150. Klocke, J. L. et al. Single-shot sub-microsecond mid-infrared spectroscopy on protein reactions with quantum cascade laser frequency combs. Anal. Chem. 90, 10494–10500 (2018).

    Article  CAS  PubMed  Google Scholar 

  151. Hamm, P., Kaindl, R. A. & Stenger, J. Noise suppression in femtosecond mid-infrared light sources. Opt. Lett. 25, 1798–1800 (2000).

    Article  CAS  PubMed  Google Scholar 

  152. Bredenbeck, J., Helbing, J. & Hamm, P. Continuous scanning from picoseconds to microseconds in time resolved linear and nonlinear spectroscopy. Rev. Sci. Instrum. 75, 4462–4466 (2004).

    Article  CAS  Google Scholar 

  153. Greetham, G. M. et al. A 100 kHz time-resolved multiple-probe femtosecond to second infrared absorption spectrometer. Appl. Spectrosc. 70, 645–653 (2016).

    Article  CAS  PubMed  Google Scholar 

  154. Rau, H. Spectroscopic properties of organic azo compounds. Angew. Chem. Int. Ed. Engl. 12, 224–235 (1973).

    Article  Google Scholar 

  155. Hobson, M. P. & Lasenby, A. N. The entropic prior for distributions with positive and negative values. Mon. Not. R. Astron. Soc. 298, 905–908 (1998).

    Article  Google Scholar 

  156. Kumar, A. T. N., Zhu, L., Christian, J. F., Demidov, A. A. & Champion, P. M. On the rate distribution analysis of kinetic data using the maximum entropy method: applications to myoglobin relaxation on the nanosecond and femtosecond timescales. J. Phys. Chem. B 105, 7847–7856 (2001).

    Article  CAS  Google Scholar 

  157. Lórenz-Fonfría, V. A. & Kandori, H. Transformation of time-resolved spectra to lifetime-resolved spectra by maximum entropy inversion of the Laplace transform. Appl. Spectrosc. 60, 407–417 (2006).

    Article  PubMed  Google Scholar 

  158. Shaw, D. E. et al. Atomic-level characterization of the structural dynamics of proteins. Science 330, 341–346 (2010).

    Article  CAS  PubMed  Google Scholar 

  159. Bowman, G. R., Pande, V. S. & Noe, F. An Introduction to Markov State Models (Springer, 2013).

  160. Pande, V. S., Beauchamp, K. & Bowman, G. R. Everything you wanted to know about Markov state models but were afraid to ask. Methods 52, 99–105 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  161. Prinz, J.-H. et al. Markov models of molecular kinetics: generation and validation. J. Chem. Phys. 134, 174105 (2011).

    Article  PubMed  Google Scholar 

  162. Sengupta, U. & Strodel, B. Markov models for the elucidation of allosteric regulation. Phil. Trans. R. Soc. B 373, 20170178 (2018).

    Article  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

The authors thank their co-workers in this project, in particular, B. Buchli, S. Waldauer, R. Walser, O. Zerbe, R. Pfister, K. Koziol, P. J. M. Johnson, C. Zanobini, J. Ruf and D. Buhrke, as well as the groups of G. Stock, A. Caflisch and B. Schuler for their numerous contributions. The authors also thank A. Woolley, who got them started in an initial phase of the project. The authors’ work has been supported by a European Research Council (ERC) Advanced Investigator Grant (DYNALLO) and the Swiss National Science Foundation (SNF) through the NCCR MUST and grant numbers 200021_165789/1 and 200020B_188694/1.

Author information

Authors and Affiliations

  1. Department of Chemistry, University of Zurich, Zurich, Switzerland

    Olga Bozovic, Brankica Jankovic & Peter Hamm

Authors
  1. Olga Bozovic
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Brankica Jankovic
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Peter Hamm
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

The authors contributed equally to all aspects of the article.

Corresponding author

Correspondence to Peter Hamm.

Ethics declarations

Competing interests

The authors declare no competing interests.

Additional information

Peer review information

Nature Reviews Chemistry thanks J. Wachtveitl, who co-reviewed with C. Slavov; and L. Chen for their contribution to the peer review of this work.

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Bozovic, O., Jankovic, B. & Hamm, P. Using azobenzene photocontrol to set proteins in motion. Nat Rev Chem 6, 112–124 (2022). https://doi.org/10.1038/s41570-021-00338-6

Download citation

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41570-021-00338-6

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing