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A Rotamer Library Approach to Modeling Side Chain Ensembles of the Bifunctional Spin Label RX

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Abstract

We introduce a rotamer library-based approach to modeling side chain ensembles of bifunctional spin labels. Because the bifunctional label is constrained by two attachment sites, the label is split into two monofunctional rotamers which are first attached to their respective sites, and then rejoined by a local optimization in dihedral space. We validate this method against a set of previously published experimental data using the bifunctional nitroxide spin label, RX. This method is relatively fast and can readily be used for both experimental analysis and protein modeling, providing significant advantages over modeling bifunctional labels with molecular dynamics simulations. Use of bifunctional labels for site-directed spin labeling (SDSL) electron paramagnetic resonance (EPR) spectroscopy dramatically reduces label mobility, which can significantly improve resolution of small changes in protein backbone structure and dynamics. Coupling the use of bifunctional labels with side chain modeling methods allows for improved quantitative application of experimental SDSL EPR data to protein modeling.

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Availability of Data and Materials

An implementation of the bifunctional label modeling method is available free and open source through chiLife (https://github.com/StollLab/chiLife). The rotamer libraries developed for RX are available on the chiLife_rotlibs GitHub repository (https://github.com/StollLab/chiLife_rotlibs).

References

  1. W.L. Hubbell, D.S. Cafiso, C. Altenbach, Nat. Struct. Mol. Biol. 7, 735 (2000)

    CAS  Google Scholar 

  2. W.L. Hubbell, C. Altenbach, Curr. Opin. Struct. Biol. 4, 566 (1994)

    CAS  Google Scholar 

  3. C.J. López, S. Oga, W.L. Hubbell, Biochemistry 51, 6568 (2012)

    PubMed  Google Scholar 

  4. C. Altenbach, S.L. Flitsch, H.G. Khorana, W.L. Hubbell, Biochemistry 28, 7806 (1989)

    CAS  PubMed  Google Scholar 

  5. C. Altenbach, T. Marti, H. Khorana, W. Hubbell, Science 248, 1088 (1990)

    CAS  PubMed  Google Scholar 

  6. C. Altenbach, W. Froncisz, R. Hemker, H. Mchaourab, W.L. Hubbell, Biophys J. 89, 2103 (2005)

    CAS  PubMed  PubMed Central  Google Scholar 

  7. L. Columbus, W.L. Hubbell, Trends Biochem. Sci. 27, 288 (2002)

    CAS  PubMed  Google Scholar 

  8. C. Altenbach, D. Greenhalgh, H. Khorana, W. Hubbell, Proc. Natl. Acad. Sci. USA 91, 1667 (1994)

    CAS  PubMed  PubMed Central  Google Scholar 

  9. C. Altenbach, K.-J. Oh, R.J. Trabanino, K. Hideg, W.L. Hubbell, Biochemistry 40, 15471 (2001)

    CAS  PubMed  Google Scholar 

  10. A.D. Milov, A.G. Maryasov, Y.D. Tsvetkov, Appl. Magn. Reson. 15, 107 (1998)

    CAS  Google Scholar 

  11. A.D. Milov, A.B. Ponomarev, Yu.D. Tsvetkov, Chem. Phys. Lett. 110, 67 (1984)

    CAS  Google Scholar 

  12. M. Pannier, S. Veit, A. Godt, G. Jeschke, H.W. Spiess, J. Magn. Reson. 142, 331 (2000)

    CAS  PubMed  Google Scholar 

  13. G. Jeschke, Y. Polyhach, Phys. Chem. Chem. Phys. 9, 1895 (2007)

    CAS  PubMed  Google Scholar 

  14. G. Jeschke, A. Koch, U. Jonas, A. Godt, J. Mag. Res. 155, 72 (2002)

    CAS  Google Scholar 

  15. S. G. Worswick, J. A. Spencer, G. Jeschke, I. Kuprov, Sci. Adv. 4, eaat5218 (2018).

  16. S.M. Islam, R.A. Stein, H.S. McHaourab, B. Roux, J. Phys. Chem. B 117, 4740 (2013)

    CAS  PubMed  PubMed Central  Google Scholar 

  17. P. Sompornpisut, B. Roux, E. Perozo, Biophys. J. 95, 5349 (2008)

    CAS  PubMed  PubMed Central  Google Scholar 

  18. R. Shen, W. Han, G. Fiorin, S.M. Islam, K. Schulten, B. Roux, PLoS Comput. Biol. 11, e1004368 (2015)

    PubMed  PubMed Central  Google Scholar 

  19. K. Kazmier, S. Sharma, M. Quick, S.M. Islam, B. Roux, H. Weinstein, J.A. Javitch, H.S. Mchaourab, Nat. Struct. Mol. Biol. 21, 472 (2014)

    CAS  PubMed  PubMed Central  Google Scholar 

  20. U. Krug, N.S. Alexander, R.A. Stein, A. Keim, H.S. Mchaourab, N. Sträter, J. Meiler, Structure 24, 43 (2016)

    CAS  PubMed  Google Scholar 

  21. M.H. Tessmer, S.A. DeCero, D. del Alamo, M.O. Riegert, J. Meiler, D.W. Frank, J.B. Feix, Sci. Rep. 10, 19700 (2020)

    CAS  PubMed  PubMed Central  Google Scholar 

  22. D. del Alamo, L. DeSousa, R.M. Nair, S. Rahman, J. Meiler, H.S. Mchaourab, Proc. Natl. Acad. Sci. USA 119, e2206129119 (2022)

    PubMed  PubMed Central  Google Scholar 

  23. D. del Alamo, K.L. Jagessar, J. Meiler, H.S. Mchaourab, PLoS Comput. Biol 17, e1009107 (2021)

    PubMed  PubMed Central  Google Scholar 

  24. D. Sala, D. del Alamo, H.S. Mchaourab, J. Meiler, Structure 30, 1157 (2022)

    CAS  PubMed  PubMed Central  Google Scholar 

  25. E.G.B. Evans, J.L.W. Morgan, F. DiMaio, W.N. Zagotta, S. Stoll, Proc. Natl. Acad. Sci. USA 20, 10839 (2020)

    Google Scholar 

  26. M.C. Puljung, H.A. DeBerg, W.N. Zagotta, S. Stoll, Proc. Natl. Acad. Sci. USA 111, 9816 (2014)

    CAS  PubMed  PubMed Central  Google Scholar 

  27. J.M. Hays, M.K. Kieber, J.Z. Li, J. Han, L. Columbus, P.M. Kasson, Angew. Chem. Int. Ed. 57, 17110 (2018)

    CAS  Google Scholar 

  28. J.M. Hays, D.S. Cafiso, P.M. Kasson, J. Phys. Chem. Lett. 10, 3410 (2019)

    CAS  PubMed  PubMed Central  Google Scholar 

  29. G. Jeschke, Protein Sci. 30, 125 (2020)

    PubMed  PubMed Central  Google Scholar 

  30. M.H. Tessmer, D.M. Anderson, A.M. Pickrum, M.O. Riegert, R. Moretti, J. Meiler, J.B. Feix, D.W. Frank, Proc. Nat. Acad. Sci. USA 115, 525 (2018)

    CAS  PubMed  PubMed Central  Google Scholar 

  31. S. Kim, S. Brandon, Z. Zhou, C.E. Cobb, S.J. Edwards, C.W. Moth, C.S. Parry, J.A. Smith, T.P. Lybrand, E.J. Hustedt, A.H. Beth, J. Biol. Chem. 286, 20746 (2011)

    CAS  PubMed  PubMed Central  Google Scholar 

  32. S.J. Edwards, C.W. Moth, S. Kim, S. Brandon, Z. Zhou, C.E. Cobb, E.J. Hustedt, A.H. Beth, J.A. Smith, T.P. Lybrand, J. Phys. Chem. B 118, 4717 (2014)

    CAS  PubMed  PubMed Central  Google Scholar 

  33. S.-H. Liou, W. K. Myers, J. D. Oswald, D. R. Britt, D. B. Goodin, Biochemistry (2017).

  34. C. Engelhard, R.P. Diensthuber, A. Möglich, R. Bittl, Sci. Rep. 7, 1385 (2017)

    PubMed  PubMed Central  Google Scholar 

  35. D. Hilger, Y. Polyhach, E. Padan, H. Jung, G. Jeschke, Biophys. J. 93, 3675 (2007)

    CAS  PubMed  PubMed Central  Google Scholar 

  36. M. Zoltner, D.G. Norman, P.K. Fyfe, H.E. Mkami, T. Palmer, W.N. Hunter, Structure 21, 595 (2013)

    CAS  PubMed  PubMed Central  Google Scholar 

  37. R. Ward, M. Zoltner, L. Beer, E. H. Mkami, H. I. Structure (2009).

  38. D.D. Alamo, M.H. Tessmer, R.A. Stein, J.B. Feix, H.S. Mchaourab, J. Meiler, Biophys. J. 2, 366 (2019)

    Google Scholar 

  39. N. Alexander, M. Bortolus, A. Al-Mestarihi, H. Mchaourab, J. Meiler, Structure 16, 181 (2008)

    CAS  PubMed  PubMed Central  Google Scholar 

  40. A.W. Fischer, D.M. Anderson, M.H. Tessmer, D.W. Frank, J.B. Feix, J. Meiler, ACS Omega 2, 2977 (2017)

    CAS  PubMed  PubMed Central  Google Scholar 

  41. Z. Guo, D. Cascio, K. Hideg, W.L. Hubbell, Protein Sci. 17, 228 (2008)

    CAS  PubMed  PubMed Central  Google Scholar 

  42. P. Fajer, M. Fajer, M. Zawrotny, W. Yang, Methods Enzymol. (2015).

  43. T.F. Cunningham, S. Pornsuwan, W.S. Horne, S. Saxena, Protein Sci. 25, 1049 (2016)

    CAS  PubMed  PubMed Central  Google Scholar 

  44. H.S. Mchaourab, M.A. Lietzow, K. Hideg, W.L. Hubbell, Biochemistry 35, 7692 (1996)

    CAS  PubMed  Google Scholar 

  45. M.A. Lietzow, W.L. Hubbell, Biochemistry 43, 3137 (2004)

    CAS  PubMed  Google Scholar 

  46. L. Columbus, T. Kálai, J. Jekö, K. Hideg, W.L. Hubbell, Biochemistry 40, 3828 (2001)

    CAS  PubMed  Google Scholar 

  47. F. Tombolato, A. Ferrarini, J.H. Freed, J. Phys. Chem. B 110, 26248 (2006)

    CAS  PubMed  PubMed Central  Google Scholar 

  48. G. Jeschke, Prog. Nucl. Magn. Reson. Spectrosc. 72, 42 (2013)

    CAS  PubMed  Google Scholar 

  49. D.T. Warshaviak, V.V. Khramtsov, D. Cascio, C. Altenbach, W.L. Hubbell, J. Magn. Reson. 232, 53 (2013)

    PubMed Central  Google Scholar 

  50. E.R. Georgieva, A.S. Roy, V.M. Grigoryants, P.P. Borbat, K.A. Earle, C.P. Scholes, J.H. Freed, J. Magn. Reson. 216, 69 (2012)

    CAS  PubMed  PubMed Central  Google Scholar 

  51. M. Tominaga, S.R. Barbosa, E.F. Poletti, J. Zukerman-Schpector, R. Marchetto, S. Schreier, A.C.M.M. Paiva, C.R. Nakaie, Chem. Pharm. Bull. 49, 1027 (2001)

    CAS  Google Scholar 

  52. R. Marchetto, S. Schreier, C.R. Nakaie, J. Am. Chem. Soc. 115, 11042 (1993)

    CAS  Google Scholar 

  53. I. Tkach, U. Diederichsen, M. Bennati, Eur. Biophys. J. 50, 143 (2021)

    CAS  PubMed  PubMed Central  Google Scholar 

  54. N.L. Fawzi, M.R. Fleissner, N.J. Anthis, T. Kálai, K. Hideg, W.L. Hubbell, G.M. Clore, J. Biomol. NMR 51, 105 (2011)

    CAS  PubMed  PubMed Central  Google Scholar 

  55. M.R. Fleissner, M.D. Bridges, E.K. Brooks, D. Cascio, T. Kálai, K. Hideg, W.L. Hubbell, Proc. Nat. Acad. Sci. USA 108, 16241 (2011)

    CAS  PubMed  PubMed Central  Google Scholar 

  56. M.A. Stevens, J.E. McKay, J.L.S. Robinson, H.E. Mkami, G.M. Smith, D.G. Norman, Phys. Chem. Chem. Phys. 18, 5799 (2015)

    PubMed Central  Google Scholar 

  57. T.F. Cunningham, M.R. Putterman, A. Desai, W.S. Horne, S. Saxena, Angew. Chem. Int. Ed. 54, 6330 (2015)

    CAS  Google Scholar 

  58. S. Ghosh, M.J. Lawless, G.S. Rule, S. Saxena, J. Magn. Reson. 286, 163 (2018)

    CAS  PubMed  Google Scholar 

  59. Y. Polyhach, E. Bordignon, G. Jeschke, Phys. Chem. Chem. Phys. 13, 2356 (2010)

    PubMed  Google Scholar 

  60. K.N. Beasley, B.T. Sutch, M.M. Hatmal, R. Langen, P.Z. Qin, I.S. Haworth, Methods Enzymol. 563, 569 (2015)

    CAS  PubMed  Google Scholar 

  61. M.M. Hatmal, Y. Li, B.G. Hegde, P.B. Hegde, C.C. Jao, R. Langen, I.S. Haworth, Biopolymers 97, 35 (2012)

    CAS  PubMed  Google Scholar 

  62. G. Hagelueken, R. Ward, J.H. Naismith, O. Schiemann, Appl. Magn. Reson. 42, 377 (2012)

    CAS  PubMed  PubMed Central  Google Scholar 

  63. G. Hagelueken, D. Abdullin, R. Ward, O. Schiemann, Mol. Phys. 111, 2757 (2013)

    CAS  PubMed  PubMed Central  Google Scholar 

  64. G. Hagelueken, D. Abdullin, O. Schiemann, Methods Enzymol. 563, 595 (2015)

    CAS  PubMed  Google Scholar 

  65. G. Tesei, J.M. Martins, M.B.A. Kunze, Y. Wang, R. Crehuet, K. Lindorff-Larsen, Plos Comput. Biol. 17, e1008551 (2021)

    CAS  PubMed  PubMed Central  Google Scholar 

  66. J.A. Smith, S.J. Edwards, C.W. Moth, T.P. Lybrand, Biochemistry 52, 5577 (2013)

    CAS  PubMed  Google Scholar 

  67. M. Tessmer, E. R. Canarie, S. Stoll, Biophys J. (2022).

  68. S. Spicher, D. Abdullin, S. Grimme, O. Schiemann, Phys. Chem. Chem. Phys. 22, 24282 (2020)

    CAS  PubMed  Google Scholar 

  69. Y. Qi, J. Lee, X. Cheng, R. Shen, S. M. Islam, B. Roux, W. Im, J. Comput. Chem. 415 (2020).

  70. D. Sezer, J.H. Freed, B. Roux, J. Phys. Chem. B 112, 5755 (2008)

    CAS  PubMed  PubMed Central  Google Scholar 

  71. S. Ghosh, S. Saxena, G. Jeschke, Appl. Magn. Reson. 49, 1281 (2018)

    CAS  Google Scholar 

  72. S.M. Islam, B. Roux, J. Phys. Chem. B 119, 3901 (2015)

    CAS  PubMed  PubMed Central  Google Scholar 

  73. X. Bogetti, S. Ghosh, A.G. Jarvi, J. Wang, S. Saxena, J. Phys. Chem. B 124, 2788 (2020)

    CAS  PubMed  Google Scholar 

  74. A.A. Canutescu, R.L. Dunbrack, Protein Sci 12, 963 (2003)

    CAS  PubMed  PubMed Central  Google Scholar 

  75. R.M. Fine, H. Wang, P.S. Shenkin, D.L. Yarmush, C. Levinthal, Proteins 1, 342 (1986)

    CAS  PubMed  Google Scholar 

  76. P.S. Shenkin, D.L. Yarmush, R.M. Fine, H. Wang, C. Levinthal, Biopolymers 26, 2053 (1987)

    CAS  PubMed  Google Scholar 

  77. T. Lazaridis, M. Karplus, Proteins 35, 133 (1999)

    CAS  PubMed  Google Scholar 

  78. G. Jeschke, Protein Sci. 27, 76 (2018)

    CAS  PubMed  Google Scholar 

  79. P. Pracht, F. Bohle, S. Grimme, Phys. Chem. Chem. Phys. 22, 7169 (2020)

    CAS  PubMed  Google Scholar 

  80. S. Jo, X. Cheng, J. Lee, S. Kim, S.-J.J. Park, D.S. Patel, A.H. Beaven, K.I. Lee, H. Rui, S. Park, H.S. Lee, B. Roux, A.D. MacKerell, J.B. Klauda, Y. Qi, W. Im, J. Comput. Chem. 38, 1114 (2017)

    CAS  PubMed  Google Scholar 

  81. J. Lee, X. Cheng, J.M. Swails, M. Yeom, P.K. Eastman, J.A. Lemkul, S. Wei, J. Buckner, J. Jeong, Y. Qi, S. Jo, V.S. Pande, D.A. Case, C.L. Brooks, A.D. MacKerell, J.B. Klauda, W. Im, J. Chem. Theory Comput. 12, 405 (2016)

    CAS  PubMed  Google Scholar 

  82. S. Spicher, S. Grimme, J. Phys. Chem. Lett. 6606 (2020).

  83. S. Spicher, S. Grimme, Angew. Chem. Int. Ed. 15665 (2020).

  84. M.H. Tessmer, S. Stoll, PLos Comput. Biol. 19, e1010834 (2023)

    CAS  PubMed  PubMed Central  Google Scholar 

  85. L.F. Ibáñez, G. Jeschke, S. Stoll, Magn. Reson. 1, 209 (2020)

    Google Scholar 

  86. G. Jeschke, V. Chechik, P. Ionita, A. Godt, H. Zimmermann, J. Banham, C. Timmel, D. Hilger, H. Jung, Appl. Magn. Reson. 30, 473 (2006)

    CAS  Google Scholar 

  87. J. Keeley, T. Choudhury, L. Galazzo, E. Bordignon, A. Feintuch, D. Goldfarb, H. Russell, M.J. Taylor, J.E. Lovett, A. Eggeling, L.F. Ibáñez, K. Keller, M. Yulikov, G. Jeschke, I. Kuprov, J. Magn. Reson. 338, 107186 (2022)

    CAS  PubMed  Google Scholar 

  88. L. Fábregas-Ibáñez, G. Jeschke, S. Stoll, J. Magn. Reson. 339, 107218 (2022)

    PubMed  Google Scholar 

  89. T.H. Edwards, S. Stoll, J. Magn. Reson. 288, 58 (2018)

    CAS  PubMed  PubMed Central  Google Scholar 

  90. L.H. Weaver, B.W. Matthews, J. Mol. Biol. 193, 189 (1987)

    CAS  PubMed  Google Scholar 

  91. J.C. Sacchettini, J.I. Gordon, L.J. Banaszak, J. Mol. Biol. 208, 327 (1989)

    CAS  PubMed  Google Scholar 

  92. Y.-J. Park, K.B. Sudhoff, A.J. Andrews, L.A. Stargell, K. Luger, Nat. Struct. Mol. Biol. 15, 957 (2008)

    CAS  PubMed  PubMed Central  Google Scholar 

  93. T. Hirayama, M. Taki, M. Nakamura, T. Arata, Y. Yamamoto, Chem. Lett. 35, 834 (2006)

    CAS  Google Scholar 

  94. K. Walczewska-Szewc, B. Corry, Phys. Chem. Chem. Phys. 16, 18949 (2014)

    CAS  PubMed  Google Scholar 

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Acknowledgements

The research presented here was supported by the National Institute of Health Grant R01 GM125753 (SS). T4L and riFABP DEER data were recorded by the Hubbell group (UCLA) and generously provided by Michael Bridges. The Vps75 DEER data were recorded by the groups of David G. Norman (University of Dundee) and Graham M. Smith (University of St. Andrews) and generously provided by Hassane El Mkami. High-performance computing was performed through the Hyak computing cluster at the University of Washington.

Funding

Funded by National Institute of Health grant R01 GM125753 (SS).

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  1. Department of Chemistry, University of Washington, Seattle, WA, 98195, USA

    Maxx H. Tessmer & Stefan Stoll

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  1. Maxx H. Tessmer
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The modeling methodology and experimental design were developed by MHT and SS. The software was implemented by MHT. The original draft was prepared by MHT and edited by MHT and SS.

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Correspondence to Stefan Stoll.

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Tessmer, M.H., Stoll, S. A Rotamer Library Approach to Modeling Side Chain Ensembles of the Bifunctional Spin Label RX. Appl Magn Reson 55, 127–140 (2024). https://doi.org/10.1007/s00723-023-01576-1

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