Advertisement

Joint X-ray/NMR structure refinement of multidomain/multisubunit systems

  • Azzurra Carlon
  • Enrico Ravera
  • Giacomo Parigi
  • Garib N. Murshudov
  • Claudio Luchinat
Article

Abstract

Data integration in structural biology has become a paradigm for the characterization of biomolecular systems, and it is now accepted that combining different techniques can fill the gaps in each other’s blind spots. In this frame, one of the combinations, which we have implemented in REFMAC-NMR, is residual dipolar couplings from NMR together with experimental data from X-ray diffraction. The first are exquisitely sensitive to the local details but does not give any information about overall shape, whereas the latter encodes more the information about the overall shape but at the same time tends to miss the local details even at the highest resolutions. Once crystals are obtained, it is often rather easy to obtain a complete X-ray dataset, however it is time-consuming to obtain an exhaustive NMR dataset. Here, we discuss the effect of including a-priori knowledge on the properties of the system to reduce the number of experimental data needed to obtain a more complete picture. We thus introduce a set of new features of REFMAC-NMR that allow for improved handling of RDC data for multidomain proteins and multisubunit biomolecular complexes, and encompasses the use of pseudo-contact shifts as an additional source of NMR-based information. The new feature may either help in improving the refinement, or assist in spotting differences between the crystal and the solution data. We show three different examples where NMR and X-ray data can be reconciled to a unique structural model without invoking mobility.

Keywords

Structure refinement Residual dipolar couplings Integrated structural biology REFMAC 

Notes

Acknowledgements

The support from Fondazione Cassa di Risparmio di Firenze, MIUR PRIN 2012SK7ASN, the European Commission projects iNEXT No. 653706, West-Life No. 675858, EMBO ASTF 620–2015, and Instruct-ERIC, a Landmark ESFRI project, and specifically the CERM/CIRMMP Italy center, is acknowledged. GNM is funded by Medical Research Council (Grant No. MC_UP_A025_1012). Extensive and frank discussions with Christian Griesinger on how much information on dynamics can be reliably extracted from the NMR data of CaM-IQ are acknowledged.

Supplementary material

10858_2018_212_MOESM1_ESM.docx (3.2 mb)
Supplementary material 1 (DOCX 3299 KB)

References

  1. Andrałojć W, Luchinat C, Parigi G, Ravera E (2014) Exploring regions of conformational space occupied by two-domain proteins. J Phys Chem B 118:10576–10587.  https://doi.org/10.1021/jp504820w CrossRefGoogle Scholar
  2. Andrałojć W, Berlin K, Fushman D et al (2015) Information content of long-range NMR data for the characterization of conformational heterogeneity. J Biomol NMR 62:353–371.  https://doi.org/10.1007/s10858-015-9951-6 CrossRefGoogle Scholar
  3. Andrałojć W, Hiruma Y, Liu W-M et al (2017) Identification of productive and futile encounters in an electron transfer protein complex. Proc Natl Acad Sci USA 114:E1840–E1847.  https://doi.org/10.1073/pnas.1616813114 CrossRefGoogle Scholar
  4. Banci L, Bertini I, Bren KL et al (1996) The use of pseudocontact shifts to refine solution structures of paramagnetic metalloproteins: Met80Ala cyano-cytochrome c as an example. J Biol Inorg Chem 1:117–126CrossRefGoogle Scholar
  5. Banci L, Bertini I, Cremonini MA et al (1998a) PSEUDODYANA for NMR structure calculation of paramagnetic metalloproteins using torsion angle molecular dynamics. J Biomol NMR 12:553–557CrossRefGoogle Scholar
  6. Banci L, Bertini I, Huber JG et al (1998b) Partial orientation of oxidized and reduced cytochrome b5 at high magnetic fields: magnetic susceptibility anisotropy contributions and consequences for protein solution structure determination. J Am Chem Soc 120:12903–12909.  https://doi.org/10.1021/ja981791w CrossRefGoogle Scholar
  7. Banci L, Bertini I, Cavallaro G et al (2004) Paramagnetism-based restraints for Xplor-NIH. J Biomol NMR 28:249–261CrossRefGoogle Scholar
  8. Berlin K, O’Leary DP, Fushman D (2010) Structural assembly of molecular complexes based on residual dipolar couplings. J Am Chem Soc 132:8961–8972.  https://doi.org/10.1021/ja100447p CrossRefGoogle Scholar
  9. Berlin K, Castañeda CA, Schneidman-Duhovny D et al (2013) Recovering a representative conformational ensemble from underdetermined macromolecular structural data. J Am Chem Soc 135:16595–16609.  https://doi.org/10.1021/ja4083717 CrossRefGoogle Scholar
  10. Bertini I, Donaire A, Jiménez B et al (2001) Paramagnetism-based versus classical constraints: an analysis of the solution structure of Ca Ln calbindin D9k. J Biomol NMR 21:85–98CrossRefGoogle Scholar
  11. Bertini I, Luchinat C, Parigi G (2002a) Paramagnetic constraints: an aid for quick solution structure determination of paramagnetic metalloproteins. Concepts Magn Reson 14:259–286CrossRefGoogle Scholar
  12. Bertini I, Luchinat C, Parigi G (2002b) Magnetic susceptibility in paramagnetic NMR. Prog Nucl Magn Reson Spectrosc 40:249–273CrossRefGoogle Scholar
  13. Bertini I, Del Bianco C, Gelis I et al (2004) Experimentally exploring the conformational space sampled by domain reorientation in calmodulin. Proc Natl Acad Sci USA 101:6841–6846ADSCrossRefGoogle Scholar
  14. Bertini I, Kursula P, Luchinat C et al (2009) Accurate solution structures of proteins from X-ray data and minimal set of NMR data: calmodulin peptide complexes as examples. J Am Chem Soc 131:5134–5144CrossRefGoogle Scholar
  15. Bertini I, Giachetti A, Luchinat C et al (2010) Conformational space of flexible biological macromolecules from average data. J Am Chem Soc 132:13553–13558CrossRefGoogle Scholar
  16. Brunger AT, Kuriyan J, Karplus M (1987) Crystallographic R factor refinement by molecular dynamics. Science 235:458–460ADSCrossRefGoogle Scholar
  17. Carlon A, Ravera E, Andrałojć W et al (2016a) How to tackle protein structural data from solution and solid state: an integrated approach. Prog Nucl Magn Reson Spectrosc 92–93:54–70CrossRefGoogle Scholar
  18. Carlon A, Ravera E, Hennig J et al (2016b) Improved accuracy from Joint X-ray and NMR refinement of a protein–RNA complex structure. J Am Chem Soc 138:1601–1610.  https://doi.org/10.1021/jacs.5b11598 CrossRefGoogle Scholar
  19. Cerofolini L, Fields GB, Fragai M et al (2013) Examination of matrix metalloproteinase-1 in solution: a preference for the pre-collagenolysis state. J Biol Chem 288:30659–30671.  https://doi.org/10.1074/jbc.M113.477240 CrossRefGoogle Scholar
  20. Chao JA, Williamson JR (2004) Joint X-ray and NMR refinement of the yeast L30e-mRNA complex. Structure 12:1165–1176.  https://doi.org/10.1016/j.str.2004.04.023 CrossRefGoogle Scholar
  21. Chen VB, Arendall WB, Headd JJ et al (2009) MolProbity: all-atom structure validation for macromolecular crystallography. Acta Crystallogr D 66:12–21.  https://doi.org/10.1107/S0907444909042073 CrossRefGoogle Scholar
  22. Chou JJ, Li S, Klee CB, Bax A (2001) Solution structure of Ca2+ calmodulin reveals flexible hand-like properties of its domains. Nat Struct Mol Biol 8:990–997CrossRefGoogle Scholar
  23. Clore GM (2011) Exploring sparsely populated states of macromolecules by diamagnetic and paramagnetic NMR relaxation. Protein Sci 20:229–246.  https://doi.org/10.1002/pro.576 CrossRefGoogle Scholar
  24. Cornilescu G, Marquardt J, Ottiger M, Bax A (1998) Validation of protein structure from anisotropic carbonyl chemical shifts in a dilute liquid crystalline phase. J Am Chem Soc 120:6836–6837CrossRefGoogle Scholar
  25. de Diego I, Kuper J, Bakalova N et al (2010) Molecular basis of the death-associated protein kinase-calcium/calmodulin regulator complex. Sci Signal 3:ra6–ra6.  https://doi.org/10.1126/scisignal.2000552 CrossRefGoogle Scholar
  26. Diaz-Moreno I, Diaz-Quintana A, De la Rosa MA, Ubbink M (2005) Structure of the complex between plastocyanin and cytochrome f from the cyanobacterium nostoc Sp. PCC 7119 as determined by paramagnetic NMR. J Biol Chem 280:18908–18915CrossRefGoogle Scholar
  27. Fenwick RB, van den Bedem H, Fraser JS, Wright PE (2014) Integrated description of protein dynamics from room-temperature X-ray crystallography and NMR. Proc Natl Acad Sci USA 111:E445–E454.  https://doi.org/10.1073/pnas.1323440111 ADSCrossRefGoogle Scholar
  28. Fragai M, Luchinat C, Parigi G, Ravera E (2013) Conformational freedom of metalloproteins revealed by paramagnetism-assisted NMR. Coord Chem Rev 257:2652–2667.  https://doi.org/10.1016/j.ccr.2013.02.009 CrossRefGoogle Scholar
  29. Gaponenko V, Sarma SP, Altieri AS et al (2004) Improving the accuracy of NMR structures of large proteins using pseudocontact shifts as long/range restraints. J Biomol NMR 28:205–212CrossRefGoogle Scholar
  30. Gochin M, Roder H (1995) Protein structure refinement based on paramagnetic NMR shifts. applications to wild-type and mutants forms of cytochrome c. Protein Sci 4:296–305CrossRefGoogle Scholar
  31. Hass MA, Ubbink M (2014) Structure determination of protein–protein complexes with long-range anisotropic paramagnetic NMR restraints. Curr Opin Struct Biol 24:45–53.  https://doi.org/10.1016/j.sbi.2013.11.010 CrossRefGoogle Scholar
  32. Hennig J, Militti C, Popowicz GM et al (2014) Structural basis for the assembly of the Sxl–Unr translation regulatory complex. Nature 515:287–290.  https://doi.org/10.1038/nature13693 ADSCrossRefGoogle Scholar
  33. Hoffman DW, Cameron CS, Davies C et al (1996) Ribosomal protein L9: a structure determination by the combined use of X-ray crystallography and NMR spectroscopy. J Mol Biol 264:1058–1071.  https://doi.org/10.1006/jmbi.1996.0696 CrossRefGoogle Scholar
  34. Jensen MR, Hansen DF, Ayna U et al (2006) On the use of pseudocontact shifts in the structure determination of metalloproteins. Magn Reson Chem 44:294–301CrossRefGoogle Scholar
  35. Knight MJ, Felli IC, Pierattelli R et al (2013) Magic angle spinning NMR of paramagnetic proteins. Acc Chem Res 46:2108–2116.  https://doi.org/10.1021/ar300349y CrossRefGoogle Scholar
  36. Koehler J, Meiler J (2011) Expanding the utility of NMR restraints with paramagnetic compounds: background and practical aspects. Prog Nucl Magn Reson Spectrosc 59:360–389.  https://doi.org/10.1016/j.pnmrs.2011.05.001 CrossRefGoogle Scholar
  37. Kovalevskiy O, Nicholls RA, Long F, Carlon A, Murshudov GN (2018) Overview of refinement procedures within 5: utilizing data from different sources. Acta Crystallographica Section D Struct Biol 74(3):215–227CrossRefGoogle Scholar
  38. Kurland RJ, McGarvey BR (1970) Isotropic NMR shifts in transition metal complexes: calculation of the Fermi contact and pseudocontact terms. J Magn Reson 2:286–301ADSGoogle Scholar
  39. Kursula P (2014) The many structural faces of calmodulin: a multitasking molecular jackknife. Amino Acids 46:2295–2304.  https://doi.org/10.1007/s00726-014-1795-y CrossRefGoogle Scholar
  40. Lange OF, Lakomek N-A, Farés C et al (2008) Recognition dynamics up to microseconds revealed from an RDC-derived ubiquitin ensemble in solution. Science 320:1471–1475ADSCrossRefGoogle Scholar
  41. Miller M, Lubkowski J, Mohana Rao JK et al (1996) The oligomerization domain of p53: crystal structure of the trigonal form. FEBS Lett 399:166–170.  https://doi.org/10.1016/S0014-5793(96)01231-8 CrossRefGoogle Scholar
  42. Murshudov GN, Vagin AA, Dodson EJ (1997) Refinement of macromolecular structures by the maximum-likelihood method. Acta Crystallogr D 53:240–255.  https://doi.org/10.1107/S0907444996012255 CrossRefGoogle Scholar
  43. Murshudov GN, Skubák P, Lebedev AA et al (2011) REFMAC5 for the refinement of macromolecular crystal structures. Acta Crystallogr D 67:355–367.  https://doi.org/10.1107/S0907444911001314 CrossRefGoogle Scholar
  44. Nitsche C, Otting G (2017) Pseudocontact shifts in biomolecular NMR using paramagnetic metal tags. Prog Nuclear Magn Reson Spectrosc 98–99:20–49.  https://doi.org/10.1016/j.pnmrs.2016.11.001 CrossRefGoogle Scholar
  45. Pintacuda G, John M, Su X-C, Otting G (2007) NMR structure determination of protein–ligand complexes by lanthanide labeling. Acc Chem Res 40:206–212.  https://doi.org/10.1021/ar050087z CrossRefGoogle Scholar
  46. Ravera E, Parigi G, Luchinat C (2017) Perspectives on paramagnetic NMR from a life sciences infrastructure. J Magn Reson 282:154–169.  https://doi.org/10.1016/j.jmr.2017.07.013 ADSCrossRefGoogle Scholar
  47. Raves ML, Doreleijer JF, Vis H et al (2001) Joint refinement as a tool for thorough comparison between NMR and X-ray data and structures of HU protein. J Biomol NMR 21:235–248CrossRefGoogle Scholar
  48. Rinaldelli M, Ravera E, Calderone V et al (2014) Simultaneous use of solution NMR and X-ray data in REFMAC5 for joint refinement/detection of structural differences. Acta Crystallogr D 70:958–967.  https://doi.org/10.1107/S1399004713034160 CrossRefGoogle Scholar
  49. Rinaldelli M, Carlon A, Ravera E et al (2015) FANTEN: a new web-based interface for the analysis of magnetic anisotropy-induced NMR data. J Biomol NMR 61:21–34.  https://doi.org/10.1007/s10858-014-9877-4 CrossRefGoogle Scholar
  50. Russo L, Maestre-Martinez M, Wolff S et al (2013) Interdomain dynamics explored by paramagnetic NMR. J Am Chem Soc 135:17111–17120.  https://doi.org/10.1021/ja408143f CrossRefGoogle Scholar
  51. Schiffer CA, Huber R, Wüthrich K, van Gunsteren WF (1994) Simultaneous refinement of the structure of BPTI against NMR data measured in solution and X-ray diffraction data measured in single crystals. J Mol Biol 241:588–599.  https://doi.org/10.1006/jmbi.1994.1533 CrossRefGoogle Scholar
  52. Schlundt A, Tants J-N, Sattler M (2017) Integrated structural biology to unravel molecular mechanisms of protein-RNA recognition. Methods 118–119:119–136.  https://doi.org/10.1016/j.ymeth.2017.03.015 CrossRefGoogle Scholar
  53. Schmitz C, Bonvin AMJJ (2011) Protein-protein HADDocking using exclusively pseudocontact shifts. J Biomol NMR 50:263–266.  https://doi.org/10.1007/s10858-011-9514-4 CrossRefGoogle Scholar
  54. Schmitz C, Vernon R, Otting G et al (2012) Protein structure determination from pseudocontact shifts using ROSETTA. J Mol Biol 416:668–677.  https://doi.org/10.1016/j.jmb.2011.12.056 CrossRefGoogle Scholar
  55. Schwieters CD, Kuszewski JJ, Tjandra N, Marius Clore G (2003) The Xplor-NIH NMR molecular structure determination package. J Magn Reson 160:65–73.  https://doi.org/10.1016/S1090-7807(02)00014-9 ADSCrossRefGoogle Scholar
  56. Shaanan B, Gronenborn A, Cohen G et al (1992) Combining experimental information from crystal and solution studies: joint X-ray and NMR refinement. Science 257:961–964.  https://doi.org/10.1126/science.1502561 ADSCrossRefGoogle Scholar
  57. Simon B, Madl T, Mackereth CD et al (2010) An efficient protocol for NMR-spectroscopy-based structure determination of protein complexes in solution. Angew Chem Int Ed Engl 49:1967–1970.  https://doi.org/10.1002/anie.200906147 CrossRefGoogle Scholar
  58. Tang C, Schwieters CD, Clore GM (2007) Open-to-closed transition in apo maltose-binding protein observed by paramagnetic NMR. Nature 449:1078–1082.  https://doi.org/10.1038/nature06232 ADSCrossRefGoogle Scholar
  59. Tang M, Sperling LJ, Berthold DA et al (2011) High-resolution membrane protein structure by joint calculations with solid-state NMR and X-ray experimental data. J Biomol NMR 51:227–233.  https://doi.org/10.1007/s10858-011-9565-6 CrossRefGoogle Scholar
  60. Tolman JR, Flanagan JM, Kennedy MA, Prestegard JH (1995) Nuclear magnetic dipole interactions in field-oriented proteins: information for structure determination in solution. Proc Natl Acad Sci USA 92:9279–9283ADSCrossRefGoogle Scholar
  61. Valafar H, Prestegard JH (2004) REDCAT: a residual dipolar coupling analysis tool. J Magn Reson 167:228–241.  https://doi.org/10.1016/j.jmr.2003.12.012 ADSCrossRefGoogle Scholar
  62. van den Bedem H, Fraser JS (2015) Integrative, dynamic structural biology at atomic resolution–it’s about time. Nat Methods 12:307–318.  https://doi.org/10.1038/nmeth.3324 CrossRefGoogle Scholar
  63. Volkov AN, Worrall JAR, Holtzmann E, Ubbink M (2006) Solution structure and dynamics of the complex between cytochrome c and cytochrome c peroxidase determined by paramagnetic NMR. Proc Natl Acad Sci USA 103:18945–18950.  https://doi.org/10.1073/pnas.0603551103 ADSCrossRefGoogle Scholar
  64. Ward AB, Sali A, Wilson IA (2013) Integrative structural biology. Science 339:913–915.  https://doi.org/10.1126/science.1228565 ADSCrossRefGoogle Scholar
  65. Winn MD, Ballard CC, Cowtan KD et al (2011) Overview of the CCP 4 suite and current developments. Acta Crystallogr Sect D 67:235–242.  https://doi.org/10.1107/S0907444910045749 CrossRefGoogle Scholar
  66. Zweckstetter M, Bax A (2002) Evaluation of uncertainty in alignment tensors obtained from dipolar couplings. J Biomol NMR 23:127–137CrossRefGoogle Scholar

Copyright information

© Springer Nature B.V. 2018

Authors and Affiliations

  1. 1.Magnetic Resonance Center (CERM) and Interuniversity Consortium for Magnetic Resonance of Metallo Proteins (CIRMMP)Sesto FiorentinoItaly
  2. 2.Department of Chemistry “Ugo Schiff”University of FlorenceSesto FiorentinoItaly
  3. 3.MRC Laboratory for Molecular BiologyCambridgeUK

Personalised recommendations