Towards Molecular Movies of Enzymes

  • Christopher KupitzEmail author
  • Marius Schmidt


Macromolecular crystallography has been highly successful in the past 60 years as it has been the predominant method to solve macromolecular structures, with more than 100,000 protein structures determined and posted to structural databases [, (Berman et al., Nucleic Acids Res 28:235–242, 2000)]. Crystallography methods are capable of determining structures at high resolution (<1.5 Å) as demonstrated by the many structures available at this or better resolution. A central objective of structural biology is not only to solve static structures but to also observe their associated dynamics to infer and explore their functions. To examine reactions that occur in biological macromolecules, time-resolved methods are required. In time-resolved crystallography, a reaction is triggered inside a crystal and the progress of this reaction is then probed by short but highly intense X-ray pulses, shorter than both the dynamics studied and the reaction trigger. Time-resolved crystallographic experiments have been successfully carried out at synchrotron X-ray sources (Moffat, Annu Rev Biophys Biophys Chem 18:309–332, 1989; Moffat, Chem Rev 101:1569–1581, 2001; Schmidt, Synchrotron Radiat News 28:25–30, 2015). Mainly cyclic reversible, and light-activated reactions were examined. Irreversible (single path) reactions, for example those catalyzed by enzymes, remain difficult to investigate. The initiation of a reaction by adding a substrate or ligand to protein crystals remains a challenge, which prevents routine applications. The arrival of X-ray free electron lasers and micro-focus synchrotron beamlines, with their intense X-ray pulses, permit the use of significantly smaller crystals. With small crystals faster diffusion times are achieved which allow for straightforward investigations of these reactions. Several successful experiments have already been reported which show how the structures of transiently occupied intermediates and their dynamics can be investigated at room temperature in real time. In this chapter we will discuss the experimental setup, feasibility, and potential impact of the new facilities on the field of enzymology.


  1. 1.
    DePristo, M. A., de Bakker, P. I., & Blundell, T. L. (2004). Heterogeneity and inaccuracy in protein structures solved by X-ray crystallography. Structure, 12, 831–838.PubMedCrossRefGoogle Scholar
  2. 2.
    Botha, S., Nass, K., Barends, T. R., Kabsch, W., Latz, B., Dworkowski, F., et al. (2015). Room-temperature serial crystallography at synchrotron X-ray sources using slowly flowing free-standing high-viscosity microstreams. Acta Crystallographica. Section D, Biological Crystallography, 71, 387–397.PubMedCrossRefGoogle Scholar
  3. 3.
    Frauenfelder, H., Sligar, S. G., & Wolynes, P. G. (1991). The energy landscapes and motions of proteins. Science, 254, 1598–1603.PubMedCrossRefGoogle Scholar
  4. 4.
    McCammon, J. A., & Harvey, S. C. (1987). Dynamics of proteins and nucleic acids. Cambridge, UK: Cambridge University Press. CrossRefGoogle Scholar
  5. 5.
    Steinbach, P. J., Ansari, A., Berendzen, J., Braunstein, D., Chu, K., Cowen, B. R., et al. (1991). Ligand binding to heme proteins: Connection between dynamics and function. Biochemistry, 30, 3988–4001.PubMedCrossRefGoogle Scholar
  6. 6.
    Moffat, K. (1989). Time-resolved macromolecular crystallography. Annual Review of Biophysics and Biophysical Chemistry, 18, 309–332.PubMedCrossRefGoogle Scholar
  7. 7.
    Moffat, K. (2001). Time-resolved biochemical crystallography: A mechanistic perspective. Chemical Reviews, 101, 1569–1581.PubMedCrossRefGoogle Scholar
  8. 8.
    Schmidt, M. (2008). Structure based enzyme kinetics by time-resolved X-ray crystallography. In Ultrashort laser pulses in medicine and biology. Berlin, Germany: Springer.Google Scholar
  9. 9.
    Schmidt, M. (2015). Time-resolved crystallography at X-ray free Electron lasers and synchrotron light sources. Synchrotron Radiation News, 28, 25–30.CrossRefGoogle Scholar
  10. 10.
    Stoddard, B. L. (2001). Trapping reaction intermediates in macromolecular crystals for structural analyses. Methods, 24, 125–138.PubMedCrossRefGoogle Scholar
  11. 11.
    Bourgeois, D., & Weik, M. (2009). Kinetic protein crystallography: A tool to watch proteins in action. Crystallography Reviews, 15, 87–118.CrossRefGoogle Scholar
  12. 12.
    Flint, A. J., Tiganis, T., Barford, D., & Tonks, N. K. (1997). Development of “substrate-trapping” mutants to identify physiological substrates of protein tyrosine phosphatases. Proceedings of the National Academy of Sciences of the United States of America, 94, 1680–1685.PubMedPubMedCentralCrossRefGoogle Scholar
  13. 13.
    Buch, I., Giorgino, T., & De Fabritiis, G. (2011). Complete reconstruction of an enzyme-inhibitor binding process by molecular dynamics simulations. Proceedings of the National Academy of Sciences of the United States of America, 108, 10184–10189.PubMedPubMedCentralCrossRefGoogle Scholar
  14. 14.
    Schmidt, M., & Saldin, D. K. (2014). Enzyme transient state kinetics in crystal and solution from the perspective of a time-resolved crystallographer. Structural Dynamics, 1, 1–14.CrossRefGoogle Scholar
  15. 15.
    Steinfeld, J. I., Francisco, J. S., & Hase, W. L. (1985). Chemical kinetics and dynamics. Upper Saddle River, NJ: Prentience Hall.Google Scholar
  16. 16.
    Schmidt, M., Srajer, V., Henning, R., Ihee, H., Purwar, N., Tenboer, J., et al. (2013). Protein energy landscapes determined by five-dimensional crystallography. Acta Crystallographica Section D, 69, 2534–2542.CrossRefGoogle Scholar
  17. 17.
    Rajagopal, S., Anderson, S., Srajer, V., Schmidt, M., Pahl, R., & Moffat, K. (2005). A structural pathway for signaling in the E46Q mutant of photoactive yellow protein. Structure, 13, 55–63.PubMedCrossRefGoogle Scholar
  18. 18.
    Schmidt, M., Rajagopal, S., Ren, Z., & Moffat, K. (2003). Application of singular value decomposition to the analysis of time-resolved macromolecular X-ray data. Biophysical Journal, 84, 2112–2129.PubMedPubMedCentralCrossRefGoogle Scholar
  19. 19.
    Ihee, H., Rajagopal, S., Srajer, V., Pahl, R., Anderson, S., Schmidt, M., et al. (2005). Visualizing reaction pathways in photoactive yellow protein from nanoseconds to seconds. Proceedings of the National Academy of Sciences of the United States of America, 102, 7145–7150.PubMedPubMedCentralCrossRefGoogle Scholar
  20. 20.
    Jung, Y. O., Lee, J. H., Kim, J., Schmidt, M., Moffat, K., Srajer, V., et al. (2013). Volume-conserving trans-cis isomerization pathways in photoactive yellow protein visualized by picosecond X-ray crystallography. Nature Chemistry, 5, 212–220.PubMedPubMedCentralCrossRefGoogle Scholar
  21. 21.
    Knapp, J. E., Pahl, R., Srajer, V., & Royer, W. E., Jr. (2006). Allosteric action in real time: Time-resolved crystallographic studies of a cooperative dimeric hemoglobin. Proceedings of the National Academy of Sciences of the United States of America, 103, 7649–7654.PubMedPubMedCentralCrossRefGoogle Scholar
  22. 22.
    Schmidt, M., Nienhaus, K., Pahl, R., Krasselt, A., Anderson, S., Parak, F., et al. (2005). Ligand migration pathway and protein dynamics in myoglobin: A time-resolved crystallographic study on L29W MbCO. Proceedings of the National Academy of Sciences of the United States of America, 102, 11704–11709.PubMedPubMedCentralCrossRefGoogle Scholar
  23. 23.
    Schotte, F., Cho, H. S., Kaila, V. R., Kamikubo, H., Dashdorj, N., Henry, E. R., et al. (2012). Watching a signaling protein function in real time via 100-ps time-resolved Laue crystallography. Proceedings of the National Academy of Sciences of the United States of America, 109, 19256–19261.PubMedPubMedCentralCrossRefGoogle Scholar
  24. 24.
    Schotte, F., Lim, M., Jackson, T. A., Smirnov, A. V., Soman, J., Olson, J. S., et al. (2003). Watching a protein as it functions with 150-ps time-resolved X-ray crystallography. Science, 300, 1944–1947.PubMedCrossRefGoogle Scholar
  25. 25.
    Srajer, V., Ren, Z., Teng, T. Y., Schmidt, M., Ursby, T., Bourgeois, D., et al. (2001). Protein conformational relaxation and ligand migration in myoglobin: A nanosecond to millisecond molecular movie from time-resolved Laue X-ray diffraction. Biochemistry, 40, 13802–13815.PubMedCrossRefGoogle Scholar
  26. 26.
    Srajer, V., Teng, T. Y., Ursby, T., Pradervand, C., Ren, Z., Adachi, S., et al. (1996). Photolysis of the carbon monoxide complex of myoglobin: Nanosecond time-resolved crystallography. Science, 274, 1726–1729.PubMedCrossRefGoogle Scholar
  27. 27.
    Tripathi, S., Srajer, V., Purwar, N., Henning, R., & Schmidt, M. (2012). pH dependence of the photoactive yellow protein photocycle investigated by time-resolved crystallography. Biophysical Journal, 102, 325–332.PubMedPubMedCentralCrossRefGoogle Scholar
  28. 28.
    Boutet, S., Lomb, L., Williams, G. J., Barends, T. R., Aquila, A., Doak, R. B., et al. (2012). High-resolution protein structure determination by serial femtosecond crystallography. Science, 337(6092), 362–364.PubMedPubMedCentralCrossRefGoogle Scholar
  29. 29.
    Chapman, H. N., Fromme, P., Barty, A., White, T. A., Kirian, R. A., Aquila, A., et al. (2011). Femtosecond X-ray protein nanocrystallography. Nature, 470, 73–77.PubMedPubMedCentralCrossRefGoogle Scholar
  30. 30.
    Kupitz, C., Basu, S., Grotjohann, I., Fromme, R., Zatsepin, N. A., Rendek, K. N., et al. (2014). Serial time-resolved crystallography of photosystem II using a femtosecond X-ray laser. Nature, 513, 5.CrossRefGoogle Scholar
  31. 31.
    Pande, K., Hutchison, C. D. M., Groenhof, G., Aquila, A., Robinson, J. S., Tenboer, J., et al. (2016). Femtosecond structural dynamics drives the trans/Cis isomerization in photoactive yellow protein. Science, 352, 725–729.PubMedPubMedCentralCrossRefGoogle Scholar
  32. 32.
    Tenboer, J., Basu, S., Zatsepin, N., Pande, K., Milathianaki, D., Frank, M., et al. (2014). Time-resolved serial crystallography captures high-resolution intermediates of photoactive yellow protein. Science, 346, 1242–1246.PubMedPubMedCentralCrossRefGoogle Scholar
  33. 33.
    Neutze, R., Wouts, R., van der Spoel, D., Weckert, E., & Hajdu, J. (2000). Potential for biomolecular imaging with femtosecond X-ray pulses. Nature, 406, 752–757.PubMedCrossRefPubMedCentralGoogle Scholar
  34. 34.
    Chapman, H. N., Barty, A., Bogan, M. J., Boutet, S., Frank, M., Hau-Riege, S. P., et al. (2006). Femtosecond diffractive imaging with a soft-X-ray free-electron laser. Nature Physics, 2, 839–843.CrossRefGoogle Scholar
  35. 35.
    Aquila, A., Hunter, M. S., Doak, R. B., Kirian, R. A., Fromme, P., White, T. A., et al. (2012). Time-resolved protein nanocrystallography using an X-ray free-electron laser. Optics Express, 20, 2706–2716.PubMedPubMedCentralCrossRefGoogle Scholar
  36. 36.
    Barty, A., Caleman, C., Aquila, A., Timneanu, N., Lomb, L., White, T. A., et al. (2012). Self-terminating diffraction gates femtosecond X-ray nanocrystallography measurements. Nature Photonics, 6, 35–40.PubMedCrossRefGoogle Scholar
  37. 37.
    Parak, F. G., Achterhold, K., Croci, S., & Schmidt, M. (2007). A physical picture of protein dynamics and conformational changes. Journal of Biological Physics, 33, 371–387.PubMedCrossRefGoogle Scholar
  38. 38.
    Bourgeois, D., & Weik, M. (2005). New perspectives in kinetic protein crystallography using caged compounds. Hoboken, NJ: Wiley.Google Scholar
  39. 39.
    Calvey, G. D., Katz, A. M., Schaffer, C. B., & Pollack, L. (2016). Mixing injector enables time-resolved crystallography with high hit rate at X-ray free electron lasers. Structural Dynamics, 3, 054301.PubMedPubMedCentralCrossRefGoogle Scholar
  40. 40.
    Kupitz, C., Olmos, J., Holl, M., Tremblay, L. W., Pande, K., Pandey, S., et al. (2017). Structural enzymology using X-ray free Electron lasers. Structural Dynamics, 4, 044003.PubMedCrossRefGoogle Scholar
  41. 41.
    Hekstra, D. R., White, K. I., Socolich, M. A., Henning, R. W., Srajer, V., & Ranganathan, R. (2016). Electric-field-stimulated protein mechanics. Nature, 540, 400–405.PubMedPubMedCentralCrossRefGoogle Scholar
  42. 42.
    Spence, J. C., Weierstall, U., & Chapman, H. N. (2012). X-ray lasers for structural and dynamic biology. Reports on progress in physics. Physical Society, 75, 102601.Google Scholar
  43. 43.
    Key, J. M., Srajer, V., Pahl, R., & Moffat, K. (2004). Time-resolved crystallographic studies of the heme-based sensor protein FixL. Biophysical Journal, 86, 246a–246a.Google Scholar
  44. 44.
    Barends, T. R., Foucar, L., Ardevol, A., Nass, K., Aquila, A., Botha, S., et al. (2015). Direct observation of ultrafast collective motions in CO myoglobin upon ligand dissociation. Science, 350(6259), 445–450. PubMedCrossRefGoogle Scholar
  45. 45.
    Barty, A., Kirian, R. A., Maia, F. R. N. C., Hantke, M., Yoon, C. H., White, T. A., et al. (2014). Cheetah: Software for high-throughput reduction and analysis of serial femtosecond X-ray diffraction data. Journal of Applied Crystallography, 47, 1118–1131.PubMedPubMedCentralCrossRefGoogle Scholar
  46. 46.
    Bourgeois, D., Vallone, B., Arcovito, A., Sciara, G., Schotte, F., Anfinrud, P. A., et al. (2006). Extended subnanosecond structural dynamics of myoglobin revealed by Laue crystallography. Proceedings of the National Academy of Sciences of the United States of America, 103, 4924–4929.PubMedPubMedCentralCrossRefGoogle Scholar
  47. 47.
    Nienhaus, K., Ostermann, A., Nienhaus, G. U., Parak, F. G., & Schmidt, M. (2005). Ligand migration and protein fluctuations in myoglobin mutant L29W. Biochemistry, 44, 5095–5105.PubMedCrossRefGoogle Scholar
  48. 48.
    Schmidt, M., Graber, T., Henning, R., & Srajer, V. (2010). Five-dimensional crystallography. Acta crystallographica. Section A, Foundations of crystallography, 66, 198–206.PubMedPubMedCentralCrossRefGoogle Scholar
  49. 49.
    Schmidt, M., Ihee, H., Pahl, R., & Srajer, V. (2005). Protein-ligand interaction probed by time-resolved crystallography. Methods in Molecular Biology, 305, 115–154.PubMedGoogle Scholar
  50. 50.
    Van Brederode, M. E., Hoff, W. D., Van Stokkum, I. H., Groot, M. L., & Hellingwerf, K. J. (1996). Protein folding thermodynamics applied to the photocycle of the photoactive yellow protein. Biophysical Journal, 71, 365–380.PubMedPubMedCentralCrossRefGoogle Scholar
  51. 51.
    Hutchison, C. D. M., Tenboer, J., Kupitz, C., Moffat, K., Schmidt, M., & van Thor, J. J. (2016). Photocycle populations with femtosecond excitation of crystalline photoactive yellow protein. Chemical Physics Letters, 654, 63–71.CrossRefGoogle Scholar
  52. 52.
    Lincoln, C. N., Fitzpatrick, A. E., & van Thor, J. J. (2012). Photoisomerisation quantum yield and non-linear cross-sections with femtosecond excitation of the photoactive yellow protein. Physical Chemistry Chemical Physics: PCCP, 14, 15752–15764.PubMedCrossRefGoogle Scholar
  53. 53.
    Bourgeois, D., & Royant, A. (2005). Advances in kinetic protein crystallography. Current Opinion in Structural Biology, 15, 538–547.PubMedCrossRefGoogle Scholar
  54. 54.
    Pelliccioli, A. P., & Wirz, J. (2002). Photoremovable protecting groups: Reaction mechanisms and applications. Photochemical & Photobiological Sciences, 1, 441–458.CrossRefGoogle Scholar
  55. 55.
    Goelder, M., & Givens, R. (2005). Dynamic studies in biology: Phototriggers, photoswitches and caged biomolecules. Hoboken, NJ: Wiley.CrossRefGoogle Scholar
  56. 56.
    Schlichting, I., Almo, S. C., Rapp, G., Wilson, K., Petratos, K., Lentfer, A., et al. (1990). Time-resolved X-ray crystallographic study of the conformational change in ha-Ras P21 protein on Gtp hydrolysis. Nature, 345, 309–315.CrossRefGoogle Scholar
  57. 57.
    Stoddard, B. L., Cohen, B. E., Brubaker, M., Mesecar, A. D., & Koshland, D. E., Jr. (1998). Millisecond Laue structures of an enzyme-product complex using photocaged substrate analogs. Nature Structural Biology, 5, 891–897.PubMedCrossRefGoogle Scholar
  58. 58.
    Moglich, A., & Hegemann, P. (2013). Biotechnology: Programming genomes with light. Nature, 500, 406–408.PubMedCrossRefGoogle Scholar
  59. 59.
    Moglich, A., & Moffat, K. (2010). Engineered photoreceptors as novel optogenetic tools. Photochemical & Photobiological Sciences, 9, 1286–1300.CrossRefGoogle Scholar
  60. 60.
    Crosson, S., & Moffat, K. (2002). Photoexcited structure of a plant photoreceptor domain reveals a light-driven molecular switch. The Plant Cell, 14, 1067–1075.PubMedPubMedCentralCrossRefGoogle Scholar
  61. 61.
    Geremia, S., Campagnolo, M., Demitri, N., & Johnson, L. N. (2006). Simulation of diffusion time of small molecules in protein crystals. Structure, 14, 393–400.PubMedCrossRefGoogle Scholar
  62. 62.
    Hajdu, J., Acharya, K. R., Stuart, D. I., Mclaughlin, P. J., Barford, D., Oikonomakos, N. G., et al. (1987). Catalysis in the crystal - synchrotron radiation studies with glycogen phosphorylase-B. The EMBO Journal, 6, 539–546.PubMedPubMedCentralCrossRefGoogle Scholar
  63. 63.
    Sjogren, T., Svensson-Ek, M., Hajdu, J., & Brzezinski, P. (2000). Proton-coupled structural changes upon binding of carbon monoxide to cytochrome cd1: A combined flash photolysis and X-ray crystallography study. Biochemistry, 39, 10967–10974.PubMedCrossRefGoogle Scholar
  64. 64.
    Sluyterman, L. A., & de Graaf, M. J. (1969). The activity of papain in the crystalline state. Biochimica et Biophysica Acta, 171, 277–287.PubMedCrossRefGoogle Scholar
  65. 65.
    Kim, T. H., Mehrabi, P., Ren, Z., Sljoka, A., Ing, C., Bezginov, A., et al. (2017). The role of dimer asymmetry and protomer dynamics in enzyme catalysis. Science, 355, 28104837.CrossRefGoogle Scholar
  66. 66.
    Kurisu, G., Sugimoto, A., Kai, Y., & Harada, S. (1997). A flow cell suitable for time-resolved X-ray crystallography by the Laue method. Journal of Applied Crystallography, 30, 555–556.CrossRefGoogle Scholar
  67. 67.
    Petsko, G. A. (1985). Diffraction methods for biological macromolecules. Flow cell construction and use. Methods in Enzymology, 114, 141–146.PubMedCrossRefGoogle Scholar
  68. 68.
    Chupas, P. J., Chapman, K. W., Kurtz, C., Hanson, J. C., Lee, P. L., & Grey, C. P. (2008). A versatile sample-environment cell for non-ambient X-ray scattering experiments. Journal of Applied Crystallography, 41, 822–824.CrossRefGoogle Scholar
  69. 69.
    Schmidt, M. (2013). Mix and inject, reaction initiation by diffusion for time-resolved macromolecular crystallography. Advances on Condensed Matter Physics, 2013, 1–10.CrossRefGoogle Scholar
  70. 70.
    Purwar, N., McGarry, J. M., Kostera, J., Pacheco, A. A., & Schmidt, M. (2011). Interaction of nitric oxide with catalase: Structural and kinetic analysis. Biochemistry, 50, 4491–4503.PubMedPubMedCentralCrossRefGoogle Scholar
  71. 71.
    Weierstall, U., Spence, J. C., & Doak, R. B. (2012). Injector for scattering measurements on fully solvated biospecies. The Review of Scientific Instruments, 83, 035108.PubMedCrossRefGoogle Scholar
  72. 72.
    Nogly, P., James, D., Wang, D., White, T. A., Zatsepin, N., Shilova, A., et al. (2015). Lipidic cubic phase serial millisecond crystallography using synchrotron radiation. IUCrJ, 2, 168–176.PubMedPubMedCentralCrossRefGoogle Scholar
  73. 73.
    Weierstall, U., James, D., Wang, C., White, T. A., Wang, D., Liu, W., et al. (2014). Lipidic cubic phase injector facilitates membrane protein serial femtosecond crystallography. Nature Communications, 5, 3309.PubMedPubMedCentralCrossRefGoogle Scholar
  74. 74.
    Sierra, R. G., Gati, C., Laksmono, H., Dao, E. H., Gul, S., Fuller, F., et al. (2016). Concentric-flow electrokinetic injector enables serial crystallography of ribosome and photosystem II. Nature Methods, 13, 59–62.PubMedCrossRefGoogle Scholar
  75. 75.
    Sierra, R. G., Laksmono, H., Kern, J., Tran, R., Hattne, J., Alonso-Mori, R., et al. (2012). Nanoflow electrospinning serial femtosecond crystallography. Acta Crystallographica. Section D, Biological Crystallography, 68, 1584–1587.PubMedPubMedCentralCrossRefGoogle Scholar
  76. 76.
    Hunter, M. S., Segelke, B., Messerschmidt, M., Williams, G. J., Zatsepin, N. A., Barty, A., et al. (2014). Fixed-target protein serial microcrystallography with an X-ray free electron laser. Scientific Reports, 4, 6026.PubMedPubMedCentralCrossRefGoogle Scholar
  77. 77.
    Mueller, C., Marx, A., Epp, S. W., Zhong, Y., Kuo, A., Balo, A. R., et al. (2015). Fixed target matrix for femtosecond time-resolved and in situ serial micro-crystallography. Structural Dynamics, 2(5), 054302.PubMedPubMedCentralCrossRefGoogle Scholar
  78. 78.
    Perry, S. L., Guha, S., Pawate, A. S., Henning, R., Kosheleva, I., Srajer, V., et al. (2014). In situ serial Laue diffraction on a microfluidic crystallization device. Journal of Applied Crystallography, 47, 1975–1982.PubMedPubMedCentralCrossRefGoogle Scholar
  79. 79.
    Roedig, P., Vartiainen, I., Duman, R., Panneerselvam, S., Stube, N., Lorbeer, O., et al. (2015). A micro-patterned silicon chip as sample holder for macromolecular crystallography experiments with minimal background scattering. Scientific Reports, 5, 10451.PubMedPubMedCentralCrossRefGoogle Scholar
  80. 80.
    Roessler, C. G., Agarwal, R., Allaire, M., Alonso-Mori, R., Andi, B., Bachega, J. F., et al. (2016). Acoustic injectors for drop-on-demand serial femtosecond crystallography. Structure, 24, 631–640.PubMedPubMedCentralCrossRefGoogle Scholar
  81. 81.
    Ren, Z., Perman, B., Srajer, V., Teng, T. Y., Pradervand, C., Bourgeois, D., et al. (2001). A molecular movie at 1.8 A resolution displays the photocycle of photoactive yellow protein, a eubacterial blue-light receptor, from nanoseconds to seconds. Biochemistry, 40, 13788–13801.PubMedCrossRefGoogle Scholar
  82. 82.
    Stagno, J. R., Liu, Y., Bhandari, Y. R., Conrad, C. E., Panja, S., Swain, M., et al. (2017). Structures of riboswitch RNA reaction states by mix-and-inject XFEL serial crystallography. Nature, 541(7636), 242–246.PubMedCrossRefGoogle Scholar
  83. 83.
    Adams, P. D., Afonine, P. V., Bunkoczi, G., Chen, V. B., Davis, I. W., Echols, N., et al. (2010). PHENIX: A comprehensive python-based system for macromolecular structure solution. Acta Crystallographica. Section D, Biological Crystallography, 66, 213–221.PubMedPubMedCentralCrossRefGoogle Scholar
  84. 84.
    Brunger, A. T., & Rice, L. M. (1997). Crystallographic refinement by simulated annealing: Methods and applications. Methods in Enzymology, 277, 243–269.PubMedCrossRefGoogle Scholar
  85. 85.
    Romo, T. D., Clarage, J. B., Sorensen, D. C., & Phillips, G. N., Jr. (1995). Automatic identification of discrete substates in proteins: Singular value decomposition analysis of time-averaged crystallographic refinements. Proteins, 22, 311–321.PubMedCrossRefGoogle Scholar
  86. 86.
    Henry, E. R., & Hofrichter, J. (1992). Singular value decomposition - application to analysis of experimental-data. Methods in Enzymology, 210, 129–192.CrossRefGoogle Scholar
  87. 87.
    Emsley, P., Lohkamp, B., Scott, W. G., & Cowtan, K. (2010). Features and development of coot. Acta Crystallographica. Section D, Biological Crystallography, 66, 486–501.PubMedPubMedCentralCrossRefGoogle Scholar
  88. 88.
    Murshudov, G. N., Skubak, P., Lebedev, A. A., Pannu, N. S., Steiner, R. A., Nicholls, R. A., et al. (2011). REFMAC5 for the refinement of macromolecular crystal structures. Acta Crystallographica. Section D, Biological Crystallography, 67, 355–367.PubMedPubMedCentralCrossRefGoogle Scholar
  89. 89.
    Hugonnet, J. E., Tremblay, L. W., Boshoff, H. I., Barry, C. E., 3rd, & Blanchard, J. S. (2009). Meropenem-clavulanate is effective against extensively drug-resistant Mycobacterium tuberculosis. Science, 323, 1215–1218.PubMedPubMedCentralCrossRefGoogle Scholar
  90. 90.
    Tremblay, L. W., Hugonnet, J. E., & Blanchard, J. S. (2008). Structure of the covalent adduct formed between Mycobacterium tuberculosis beta-lactamase and clavulanate. Biochemistry, 47, 5312–5316.PubMedCrossRefGoogle Scholar
  91. 91.
    Hugonnet, J. E., & Blanchard, J. S. (2007). Irreversible inhibition of the Mycobacterium tuberculosis beta-lactamase by clavulanate. Biochemistry, 46, 11998–12004.PubMedPubMedCentralCrossRefGoogle Scholar
  92. 92.
    Boyd, D. B., & Lunn, W. H. (1979). Electronic structures of cephalosporins and penicillins. 9. Departure of a leaving group in cephalosporins. Journal of Medicinal Chemistry, 22, 778–784.PubMedCrossRefGoogle Scholar
  93. 93.
    Deponte, D. P., McKeown, J. T., Weierstall, U., Doak, R. B., & Spence, J. C. (2011). Towards ETEM serial crystallography: Electron diffraction from liquid jets. Ultramicroscopy, 111, 824–827.PubMedCrossRefGoogle Scholar
  94. 94.
    Holton, J. M., & Frankel, K. A. (2010). The minimum crystal size needed for a complete diffraction data set. Acta Crystallographica Section D, 66, 393–408.CrossRefGoogle Scholar

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Authors and Affiliations

  1. 1.Physics DepartmentUniversity of Wisconsin-MilwaukeeMilwaukeeUSA

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