Journal of Biomolecular NMR

, Volume 52, Issue 1, pp 41–56 | Cite as

VITAL NMR: using chemical shift derived secondary structure information for a limited set of amino acids to assess homology model accuracy

  • Michael C. Brothers
  • Anna E. Nesbitt
  • Michael J. Hallock
  • Sanjeewa G. Rupasinghe
  • Ming Tang
  • Jason Harris
  • Jerome Baudry
  • Mary A. Schuler
  • Chad M. Rienstra


Homology modeling is a powerful tool for predicting protein structures, whose success depends on obtaining a reasonable alignment between a given structural template and the protein sequence being analyzed. In order to leverage greater predictive power for proteins with few structural templates, we have developed a method to rank homology models based upon their compliance to secondary structure derived from experimental solid-state NMR (SSNMR) data. Such data is obtainable in a rapid manner by simple SSNMR experiments (e.g., 13C–13C 2D correlation spectra). To test our homology model scoring procedure for various amino acid labeling schemes, we generated a library of 7,474 homology models for 22 protein targets culled from the TALOS+/SPARTA+ training set of protein structures. Using subsets of amino acids that are plausibly assigned by SSNMR, we discovered that pairs of the residues Val, Ile, Thr, Ala and Leu (VITAL) emulate an ideal dataset where all residues are site specifically assigned. Scoring the models with a predicted VITAL site-specific dataset and calculating secondary structure with the Chemical Shift Index resulted in a Pearson correlation coefficient (−0.75) commensurate to the control (−0.77), where secondary structure was scored site specifically for all amino acids (ALL 20) using STRIDE. This method promises to accelerate structure procurement by SSNMR for proteins with unknown folds through guiding the selection of remotely homologous protein templates and assessing model quality.


Protein structure prediction Homology modeling Solid-state NMR spectroscopy TALOS database Chemical shift analysis 



The authors thank the National Institute of Health for funding through R01GM79530, R01GM75937, NRSA (F32 GM095344), the Ruth L. Kirschstein National Research Service Award to AEN and the Chemical Biology Interface Training Program (GM070421-06) to MCB and the Department of Homeland Security Fellowship Program to MCB, as well as Dr. Ying Li, Dr. Aleksandra Kijac, and Dr. Andrew Nieuwkoop for early assistance on this project.

Supplementary material

10858_2011_9576_MOESM1_ESM.pdf (289 kb)
Supplementary material 1 (PDF 288 kb)


  1. Alber F, Förster F, Korkin D, Topf M, Sali A (2008) Integrating diverse data for structure determination of macromolecular assemblies. Annu Rev Biochem 77:443–477. doi: 10.1146/Annurev.Biochem.77.060407.135530 CrossRefGoogle Scholar
  2. Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ (1990) Basic local alignment search tool. J Mol Biol 215(3):403–410Google Scholar
  3. Baranov VI, Morozov IY, Ortlepp SA, Spirin AS (1989) Gene-expression in a cell-free system on preparative scale. Gene 84(2):463–466CrossRefGoogle Scholar
  4. Baudry J, Rupasinghe S, Schuler MA (2006) Class-dependent sequence alignment strategy improves the structural and functional modeling of P450. Protein Eng Des Sel 19(8):345–353CrossRefGoogle Scholar
  5. Bax A, Kontaxis G, Tjandra N (2001) Dipolar couplings in macromolecular structure determination. Meth Enzymol 339:127–174CrossRefGoogle Scholar
  6. Bertini I, Bhaumik A, De Paëpe G, Griffin RG, Lelli M, Lewandowski JR, Luchinat C (2010) High-resolution solid-state NMR Structure of a 17.6 kDa protein. J Am Chem Soc 132(3):1032–1040. doi: 10.1021/Ja906426p CrossRefGoogle Scholar
  7. Bowie JU, Luthy R, Eisenberg D (1991) A method to identify protein sequences that fold into a known 3-dimensional structure. Science 253(5016):164–170ADSCrossRefGoogle Scholar
  8. Carugo O, Pongor S (2001) A normalized root-mean-square distance for comparing protein three-dimensional structures. Protein Sci 10(7):1470–1473CrossRefGoogle Scholar
  9. Castellani F, van Rossum B, Diehl A, Schubert M, Rehbein K, Oschkinat H (2002) Structure of a protein determined by solid-state magic-angle-spinning NMR spectroscopy. Nature 420(6911):98–102. doi: 10.1038/Nature01070 ADSCrossRefGoogle Scholar
  10. Chou PY, Fasman GD (1974) Prediction of protein conformation. Biochemistry 13(2):222–245CrossRefGoogle Scholar
  11. De Angelis AA, Howell SC, Nevzorov AA, Opella SJ (2006) Structure determination of a membrane protein with two trans-membrane helices in aligned phospholipid bicelles by solid-state NMR spectroscopy. J Am Chem Soc 128(37):12256–12267. doi: 10.1021/Ja063640w CrossRefGoogle Scholar
  12. Endo Y, Sawasaki T (2003) High-throughput, genome-scale protein production method based on the wheat germ cell-free expression system. Biotechnol Adv 21(8):695–713. doi: 10.1016/S0734-9750(03)00105-8 CrossRefGoogle Scholar
  13. Engelman DM, Brunger A, Cocco M, Fleming K, Gerstein M, Mackenzie K, Prestegard J, Russ W, Senes A, Zhou F (2000) Helix interactions in membrane proteins. Faseb J 14(8):A1506Google Scholar
  14. Eramian D, Eswar N, Shen MY, Sali A (2008) How well can the accuracy of comparative protein structure models be predicted? Protein Sci 17(11):1881–1893. doi: 10.1110/Ps.036061.108 CrossRefGoogle Scholar
  15. Eswar N, Marti-Renom MA, Webb B, Madhusudhan MS, Eramian D, Shen M, Pieper U, Sali A (2000) Comparative protein structure modeling with MODELLER. Curr Protoc Bioinform Suppl 15:5.6.1–5.6.30Google Scholar
  16. Fasnacht M, Zhu J, Honig B (2007) Local quality assessment in homology models using statistical potentials and support vector machines. Protein Sci 16(8):1557–1568CrossRefGoogle Scholar
  17. Fischer MW, Losonczi JA, Weaver JL, Prestegard JH (1999) Domain orientation and dynamics in multidomain proteins from residual dipolar couplings. Biochemistry 38(28):9013–9022CrossRefGoogle Scholar
  18. Forrest LR, Tang CL, Honig B (2006) On the accuracy of homology modeling and sequence alignment methods applied to membrane proteins. Biophys J 91(2):508–517. doi: 10.1529/Biophysj.106.082313 CrossRefGoogle Scholar
  19. Franks WT, Kloepper KD, Wylie BJ, Rienstra CM (2007) Four-dimensional heteronuclear correlation experiments for chemical shift assignment of solid proteins. J Biomol NMR 39(2):107–131. doi: 10.1007/S10858-007-9179-1 CrossRefGoogle Scholar
  20. Franks WT, Wylie BJ, Schmidt HLF, Nieuwkoop AJ, Mayrhofer RM, Shah GJ, Graesser DT, Rienstra CM (2008) Dipole tensor-based atomic-resolution structure determination of a nanocrystalline protein by solid-state NMR. Proc Natl Acad Sci USA 105(12):4621–4626ADSCrossRefGoogle Scholar
  21. Frericks HL, Zhou DH, Yap LL, Gennis RB, Rienstra CM (2006) Magic-angle spinning solid-state NMR of a 144 kDa membrane protein complex: E. coli cytochrome bo3 oxidase. J Biomol NMR 36(1):55–71. doi: 10.1007/S10858-006-9070-5 CrossRefGoogle Scholar
  22. Frishman D, Argos P (1997) Seventy-five percent accuracy in protein secondary structure prediction. Proteins 27(3):329–335CrossRefGoogle Scholar
  23. Grishaev A, Tugarinov V, Kay LE, Trewhella J, Bax A (2008) Refined solution structure of the 82-kDa enzyme malate synthase G from joint NMR and synchrotron SAXS restraints. J Biomol NMR 40(2):95–106. doi: 10.1007/S10858-007-9211-5 CrossRefGoogle Scholar
  24. Grzesiek S, Bax A (1993) Amino-acid type determination in the sequential assignment procedures of uniformly C-13/N-15-enriched proteins. J Biomol NMR 3(2):185–204CrossRefGoogle Scholar
  25. Hanson MA, Stevens RC (2009) Discovery of new GPCR biology: one receptor structure at a time. Structure 17(1):8–14. doi: 10.1016/J.Str.2008.12.003 CrossRefGoogle Scholar
  26. Heinig M, Frishman D (2004) STRIDE: a web server for secondary structure assignment from known atomic coordinates of proteins. Nucleic Acids Res 32:W500–W502. doi: 10.1093/Nar/Gkh429 CrossRefGoogle Scholar
  27. Hiller S, Garces RG, Malia TJ, Orekhov VY, Colombini M, Wagner G (2008) Solution structure of the integral human membrane protein VDAC-1 in detergent micelles. Science 321(5893):1206–1210. doi: 10.1126/Science.1161302 ADSCrossRefGoogle Scholar
  28. Hooft RWW, Vriend G, Sander C, Abola EE (1996) Errors in protein structures. Nature 381(6580):272ADSCrossRefGoogle Scholar
  29. Humphrey W, Dalke A, Schulten K (1996) VMD: visual molecular dynamics. J Mol Graph 14(1):33–38CrossRefGoogle Scholar
  30. Ikura M, Kay LE, Krinks M, Bax A (1991) Triple-resonance multidimensional NMR-study of calmodulin complexed with the binding domain of skeletal-muscle myosin light-chain kinase—indication of a conformational change in the central helix. Biochemistry 30(22):5498–5504CrossRefGoogle Scholar
  31. Jehle S, Rajagopal P, Bardiaux B, Markovic S, Kühne R, Stout JR, Higman VA, Klevit RE, van Rossum BJ, Oschkinat H (2010) Solid-state NMR and SAXS studies provide a structural basis for the activation of alphaB-crystallin oligomers. Nat Struct Mol Biol 17(9):1037–1042. doi: 10.1038/Nsmb.1891 CrossRefGoogle Scholar
  32. Kelly K (1999) Multiple sequence and structure refinement in MOE. Chemical Computing Group Inc.
  33. Laskowski RA, Rullmann JAC, MacArthur MW, Kaptein R, Thornton JM (1996) AQUA and PROCHECK-NMR: programs for checking the quality of protein structures solved by NMR. J Biomol NMR 8(4):477–486CrossRefGoogle Scholar
  34. Li Y, Berthold DA, Frericks HL, Gennis RB, Rienstra CM (2007) Partial 13C and 15N chemical-shift assignments of the disulfide-bond-forming enzyme DsbB by 3D magic-angle spinning NMR spectroscopy. Chembiochem 8(4):434–442. doi: 10.1002/Cbic.200600484 MATHCrossRefGoogle Scholar
  35. Li Y, Berthold DA, Gennis RB, Rienstra CM (2008) Chemical shift assignment of the transmembrane helices of DsbB, a 20-kDa integral membrane enzyme, by 3D magic-angle spinning NMR spectroscopy. Protein Sci 17(2):199–204CrossRefGoogle Scholar
  36. Lin MT, Sperling LJ, Frericks Schmidt HL, Tang M, Gennis RB, Rienstra CM (2011) A rapid and robust method for selective isotope labeling of proteins for solid-state NMR and pulsed EPR studies. Methods. doi: 10.1016/j.ymeth.2011.08.019
  37. Loquet A, Bardiaux B, Gardiennet C, Blanchet C, Baldus M, Nilges M, Malliavin T, Bockmann A (2008) 3D structure determination of the Crh protein from highly ambiguous solid-state NMR restraints. J Am Chem Soc 130(11):3579–3589. doi: 10.1021/Ja078014t CrossRefGoogle Scholar
  38. Manolikas T, Herrmann T, Meier BH (2008) Protein structure determination from 13C spin-diffusion solid-state NMR spectroscopy. J Am Chem Soc 130(12):3959–3966CrossRefGoogle Scholar
  39. Marassi FM, Opella SJ (2003) Simultaneous assignment and structure determination of a membrane protein from NMR orientational restraints. Protein Sci 12(3):403–411. doi: 10.1110/Ps.0211503 CrossRefGoogle Scholar
  40. Marti-Renom MA, Stuart AC, Fiser A, Sánchez R, Melo F, Sali A (2000) Comparative protein structure modeling of genes and genomes. Annu Rev Biophys Biomol Struct 29:291–325CrossRefGoogle Scholar
  41. Melo F, Feytmans E (1997) Novel knowledge-based mean force potential at atomic level. J Mol Biol 267(1):207–222CrossRefGoogle Scholar
  42. Mercier KA, Baran M, Ramanathan V, Revesz P, Xiao R, Montelione GT, Powers R (2006) FAST-NMR: functional annotation screening technology using NMR spectroscopy. J Am Chem Soc 128(47):15292–15299. doi: 10.1021/Ja0651759 CrossRefGoogle Scholar
  43. Mobarec JC, Sanchez R, Filizola M (2009) Modern homology modeling of G-protein coupled receptors: which structural template to use? J Medicinal Chem 52(16):5207–5216. doi: 10.1021/Jm9005252 CrossRefGoogle Scholar
  44. Monleon D, Colson K, Moseley HN, Anklin C, Oswald R, Szyperski T, Montelione GT (2002) Rapid analysis of protein backbone resonance assignments using cryogenic probes, a distributed Linux-based computing architecture, and an integrated set of spectral analysis tools. J Struc Funct Genomics 2(2):93–101CrossRefGoogle Scholar
  45. Nakashima H, Nishikawa K (1994) Discrimination of intracellular and extracellular proteins using amino-acid-composition and residue-pair frequencies. J Mol Biol 238(1):54–61CrossRefGoogle Scholar
  46. Neal S, Nip AM, Zhang HY, Wishart DS (2003) Rapid and accurate calculation of protein 1H, 13C and 15N chemical shifts. J Biomol NMR 26(3):215–240CrossRefGoogle Scholar
  47. Oberai A, Ihm Y, Kim S, Bowie JU (2006) A limited universe of membrane protein families and folds. Protein Sci 15(7):1723–1734CrossRefGoogle Scholar
  48. Pearson WR (1996) Effective protein sequence comparison. Method Enzymol 266:227–258CrossRefGoogle Scholar
  49. Pieper U, Eswar N, Webb BM, Eramian D, Kely L, Barkan DT, Carter H, Mankoo P, Karchin R, Marti-Renom MA, Davis FP, Sali A (2009) MODBASE, a database of annotated comparative protein structure models and associated resources. Nucleic Acids Res 37(Database issue):D347–D354CrossRefGoogle Scholar
  50. Raman S, Lange OF, Rossi P, Tyka M, Wang X, Aramini J, Liu GH, Ramelot TA, Eletsky A, Szyperski T, Kennedy MA, Prestegard J, Montelione GT, Baker D (2010) NMR Structure determination for larger proteins using backbone-only data. Science 327(5968):1014–1018. doi: 10.1126/Science.1183649 ADSCrossRefGoogle Scholar
  51. Randazzo A, Acklin C, Schafer BW, Heizmann CW, Chazin WJ (2001) Structural insight into human Zn2+-bound S100A2 from NMR and homology modeling. Biochem Bioph Res Co 288(2):462–467CrossRefGoogle Scholar
  52. Ray A, Lindahl E, Wallner B (2010) Model quality assessment for membrane proteins. Bioinformatics 26(24):3067–3074CrossRefGoogle Scholar
  53. Robustelli P, Kohlhoff K, Cavalli A, Vendruscolo M (2010) Using NMR chemical shifts as structural restraints in molecular dynamics simulations of proteins. Structure 18(8):923–933. doi: 10.1016/J.Str.2010.04.016 CrossRefGoogle Scholar
  54. Rost B (1999) Twilight zone of protein sequence alignments. Protein Eng 12(2):85–94MathSciNetCrossRefGoogle Scholar
  55. Sali A, Blundell TL (1993) Comparative protein modeling by satisfaction of spatial restraints. J Mol Biol 234(3):779–815CrossRefGoogle Scholar
  56. Sawasaki T, Ogasawara T, Morishita R, Endo Y (2002) A cell-free protein synthesis system for high-throughput proteomics. Proc Natl Acad Sci USA 99(23):14652–14657. doi: 10.1073/Pnas.232580399 ADSCrossRefGoogle Scholar
  57. Schröder GF, Levitt M, Brunger AT (2010) Super-resolution biomolecular crystallography with low-resolution data. Nature 464(7292):1218–1222. doi: 10.1038/Nature08892 ADSCrossRefGoogle Scholar
  58. Schwarz D, Dotsch V, Bernhard F (2008) Production of membrane proteins using cell-free expression systems. Proteomics 8(19):3933–3946. doi: 10.1002/Pmic.200800171 CrossRefGoogle Scholar
  59. Senes A, Engel DE, DeGrado WF (2004) Folding of helical membrane proteins: the role of polar, GxxxG-like and proline motifs. Curr Opin Struct Biol 14(4):465–479. doi: 10.1016/J.Sbi.2004.07.007 CrossRefGoogle Scholar
  60. Shen Y, Bax A (2007) Protein backbone chemical shifts predicted from searching a database for torsion angle and sequence homology. J Biomol NMR 38(4):289–302. doi: 10.1007/S10858-007-9166-6 CrossRefGoogle Scholar
  61. Shen Y, Bax A (2010) SPARTA plus: a modest improvement in empirical NMR chemical shift prediction by means of an artificial neural network. J Biomol NMR 48(1):13–22. doi: 10.1007/S10858-010-9433-9 CrossRefGoogle Scholar
  62. Shen Y, Lange O, Delaglio F, Rossi P, Aramini JM, Liu GH, Eletsky A, Wu YB, Singarapu KK, Lemak A, Ignatchenko A, Arrowsmith CH, Szyperski T, Montelione GT, Baker D, Bax A (2008) Consistent blind protein structure generation from NMR chemical shift data. Proc Natl Acad Sci USA 105(12):4685–4690. doi: 10.1073/Pnas.0800256105 ADSCrossRefGoogle Scholar
  63. Shen Y, Delaglio F, Cornilescu G, Bax A (2009) TALOS + : a hybrid method for predicting protein backbone torsion angles from NMR chemical shifts. J Biomol NMR 44(4):213–223. doi: 10.1007/S10858-009-9333-Z CrossRefGoogle Scholar
  64. Sippl MJ (1993) Recognition of errors in 3-dimensional structures of proteins. Proteins 17(4):355–362CrossRefGoogle Scholar
  65. Spera S, Bax A (1991) Empirical correlation between protein backbone conformation and Cα and Cβ 13C nuclear-magnetic-resonance chemical-shifts. J Am Chem Soc 113(14):5490–5492CrossRefGoogle Scholar
  66. Swindells MB, Macarthur MW, Thornton JM (1995) Intrinsic phi, psi propensities of amino-acids, derived from the coil regions of known structures. Nat Struct Biol 2(7):596–603CrossRefGoogle Scholar
  67. Tang MT, Sperling LJ, Berthold DA, Schwieters CD, Nesbitt AE, Niuwkoop AJ, Gennis RB, Rienstra CM (2011) High-resolution membrane protein structure by joint calculations with solid-state NMR and X-ray experimental data. J Biol NMR. doi: 10.1007/s10858-011-9565-6
  68. Traaseth NJ, Shi L, Verardi R, Mullen DG, Barany G, Veglia G (2009) Structure and topology of monomeric phospholamban in lipid membranes determined by a hybrid solution and solid-state NMR approach. Proc Natl Acad Sci USA 106(25):10165–10170. doi: 10.1073/Pnas.0904290106 ADSCrossRefGoogle Scholar
  69. Tycko R, Hu KN (2010) A Monte Carlo/simulated annealing algorithm for sequential resonance assignment in solid state NMR of uniformly labeled proteins with magic-angle spinning. J Magn Reson 205(2):304–314ADSCrossRefGoogle Scholar
  70. Van Horn WD, Kim HJ, Ellis CD, Hadziselimovic A, Sulistijo ES, Karra MD, Tian CL, Sönnichsen FD, Sanders CR (2009) Solution nuclear magnetic resonance structure of membrane-integral diacylglycerol kinase. Science 324(5935):1726–1729. doi: 10.1126/Science.1171716 ADSCrossRefGoogle Scholar
  71. Van Melckebeke H, Wasmer C, Lange A, Eiso AB, Loquet A, Böckmann A, Meier BH (2010) Atomic-resolution three-dimensional structure of HET-s(218-289) amyloid fibrils by solid-state NMR spectroscopy. J Am Chem Soc 132(39):13765–13775. doi: 10.1021/Ja104213j CrossRefGoogle Scholar
  72. Vila JA, Aramini JM, Rossi P, Kuzin A, Su M, Seetharaman J, Xiao R, Tong L, Montelione GT, Scheraga HA (2008) Quantum chemical 13Cα chemical shift calculations for protein NMR structure determination, refinement, and validation. Proc Natl Acad Sci USA 105(38):14389–14394. doi: 10.1073/Pnas.0807105105 ADSCrossRefGoogle Scholar
  73. Wallner B, Elofsson A (2003) Can correct protein models be identified? Protein Sci 12(5):1073–1086. doi: 10.1110/Ps.0236803 CrossRefGoogle Scholar
  74. Waugh DS (1996) Genetic tools for selective labeling of proteins with alpha-15N-amino acids. J Biomol NMR 8(2):184–192CrossRefGoogle Scholar
  75. Weichenberger CX, Sippl MJ (2006) NQ-Flipper: validation and correction of asparagine/glutamine amide rotamers in protein crystal structures. Bioinformatics 22(11):1397–1398. doi: 10.1093/Bioinformatics/Bt/128 CrossRefGoogle Scholar
  76. White SH (2009) Biophysical dissection of membrane proteins. Nature 459(7245):344–346. doi: 10.1038/Nature08142 ADSCrossRefGoogle Scholar
  77. Wishart DS, Sykes BD (1994) The 13C chemical-shift index—a simple method for the identification of protein secondary structure using 13C chemical-shift data. J Biomol NMR 4(2):171–180CrossRefGoogle Scholar
  78. Wylie BJ, Schwieters CD, Oldfield E, Rienstra CM (2009) Protein structure refinement using 13Cα chemical shift tensors. J Am Chem Soc 131(3):985–992. doi: 10.1021/Ja804041p CrossRefGoogle Scholar
  79. Yang YH, Ramelot TA, McCarrick RM, Ni SS, Feldmann EA, Cort JR, Wang HA, Ciccosanti C, Jiang M, Janjua H, Acton TB, Xiao R, Everett JK, Montelione GT, Kennedy MA (2010) Combining NMR and EPR methods for homodimer protein structure determination. J Am Chem Soc 132(34):11910–11913. doi: 10.1021/Ja105080h CrossRefGoogle Scholar
  80. Yarnitzky T, Levit A, Niv MY (2010) Homology modeling of G-protein-coupled receptors with X-ray structures on the rise. Curr Opin Drug Discov Devel 13(3):317–325Google Scholar

Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  • Michael C. Brothers
    • 1
  • Anna E. Nesbitt
    • 1
  • Michael J. Hallock
    • 1
  • Sanjeewa G. Rupasinghe
    • 2
  • Ming Tang
    • 1
  • Jason Harris
    • 3
  • Jerome Baudry
    • 3
    • 4
  • Mary A. Schuler
    • 2
    • 5
  • Chad M. Rienstra
    • 1
    • 5
    • 6
  1. 1.Department of ChemistryUniversity of Illinois at Urbana-ChampaignUrbanaUSA
  2. 2.Department of Cell and Developmental BiologyUniversity of Illinois at Urbana-ChampaignUrbanaUSA
  3. 3.Department of Biochemistry, Cellular and Molecular BiologyUniversity of TennesseeKnoxvilleUSA
  4. 4.UT/ORNL Center for Molecular BiophysicsOak Ridge National LaboratoryOak RidgeUSA
  5. 5.Department of BiochemistryUniversity of Illinois at Urbana-ChampaignUrbanaUSA
  6. 6.Center for Biophysics and Computational BiologyUniversity of Illinois at Urbana-ChampaignUrbanaUSA

Personalised recommendations