Journal of Biomolecular NMR

, Volume 61, Issue 1, pp 65–72 | Cite as

J-UNIO protocol used for NMR structure determination of the 206-residue protein NP_346487.1 from Streptococcus pneumoniae TIGR4

  • Kristaps Jaudzems
  • Bill Pedrini
  • Michael Geralt
  • Pedro Serrano
  • Kurt WüthrichEmail author


The NMR structure of the 206-residue protein NP_346487.1 was determined with the J-UNIO protocol, which includes extensive automation of the structure determination. With input from three APSY-NMR experiments, UNIO-MATCH automatically yielded 77 % of the backbone assignments, which were interactively validated and extended to 97 %. With an input of the near-complete backbone assignments and three 3D heteronuclear-resolved [1H,1H]-NOESY spectra, automated side chain assignment with UNIO-ATNOS/ASCAN resulted in 77 % of the expected assignments, which was extended interactively to about 90 %. Automated NOE assignment and structure calculation with UNIO-ATNOS/CANDID in combination with CYANA was used for the structure determination of this two-domain protein. The individual domains in the NMR structure coincide closely with the crystal structure, and the NMR studies further imply that the two domains undergo restricted hinge motions relative to each other in solution. NP_346487.1 is so far the largest polypeptide chain to which the J-UNIO structure determination protocol has successfully been applied.


Putative phosphoglycolate phosphatase APSY-NMR spectroscopy NOESY Automatic data analysis Protein structure 



This work was supported by the Joint Center for Structural Genomics (JCSG) through the NIH Protein Structure Initiative (PSI) grant U54 GM094586 from the National Institute of General Medical Sciences ( Kurt Wüthrich is the Cecil H. and Ida M. Green Professor of Structural Biology at The Scripps Research Institute. We thank Dr. Ian A. Wilson for a careful reading of the manuscript.

Supplementary material

10858_2014_9886_MOESM1_ESM.tif (3.9 mb)
Supplementary material 1 (TIFF 3948 kb)


  1. Aravind L, Galperin MY, Koonin EV (1998) The catalytic domain of the P-type ATPase has the haloacid dehalogenase fold. Trends Biochem Sci 23:127–129CrossRefGoogle Scholar
  2. Burroughs AM, Allen KN, Dunaway-Mariano D, Aravind L (2006) Evolutionary genomics of the HAD superfamily: understanding the structural adaptations and catalytic diversity in a superfamily of phosphoesterases and allied enzymes. J Mol Biol 361:1003–1034CrossRefGoogle Scholar
  3. Dutta SK, Serrano P, Proudfoot A, Geralt A, Pedrini B, Herrmann T, Wüthrich K (2014) APSY-NMR for protein backbone assignment in high-throughput structural biology. J Biomol NMR. doi: 10.1007/s10858-014-9881-8
  4. Fiorito F, Herrmann T, Damberger FF, Wüthrich K (2008) Automated amino acid side-chain NMR assignment of proteins using 13C- and 15N-resolved 3D [1H,1H]-NOESY. J Biomol NMR 42:23–33CrossRefGoogle Scholar
  5. Gossert AD, Hiller S, Fernández C (2011) Automated NMR resonance assignment of large proteins for protein-ligand interaction studies. J Am Chem Soc 133:210–213CrossRefGoogle Scholar
  6. Griffin JL, Bowler MW, Baxter NJ, Leigh KN, Dannatt HR, Hounslow AM, Blackburn GM, Webster CE, Cliff MJ, Waltho JP (2012) Near attack conformers dominate β-phosphoglucomutase complexes where geometry and charge distribution reflect those of substrate. Proc Natl Acad Sci USA 109:6910–6915ADSCrossRefGoogle Scholar
  7. Güntert P, Mumenthaler C, Wüthrich K (1997) Torsion angle dynamics for NMR structure calculation with the new program DYANA. J Mol Biol 273:283–298CrossRefGoogle Scholar
  8. Herrmann T, Güntert P, Wüthrich K (2002a) Protein NMR structure determination with automated NOE-identification in the NOESY spectra using the new software ATNOS. J Biomol NMR 24:171–189CrossRefGoogle Scholar
  9. Herrmann T, Güntert P, Wüthrich K (2002b) Protein NMR structure determination with automated NOE assignment using the new software CANDID and the torsion angle dynamics algorithm DYANA. J Mol Biol 319:209–227CrossRefGoogle Scholar
  10. Hiller S, Fiorito F, Wüthrich K, Wider G (2005) Automated projection spectroscopy (APSY). Proc Natl Acad Sci USA 102:10876–10881ADSCrossRefGoogle Scholar
  11. Hiller S, Wider G, Wüthrich K (2008) APSY-NMR with proteins: practical aspects and backbone assignment. J Biomol NMR 42:179–195CrossRefGoogle Scholar
  12. Hisano T, Hata Y, Fujii T, Liu JQ, Kurihara T, Esaki N, Soda K (1996) Crystal structure of L-2-haloacid dehalogenase from Pseudomonas sp. YL. An alpha/beta hydrolase structure that is different from the alpha/beta hydrolase fold. J Biol Chem 271:20322–20330CrossRefGoogle Scholar
  13. Horst R, Wider G, Fiaux J, Bertelsen EB, Horwich AL, Wüthrich K (2006) Proton-proton Overhauser NMR spectroscopy with polypeptide chains in large structures. Proc Natl Acad Sci USA 103:15445–15450ADSCrossRefGoogle Scholar
  14. Jaudzems K, Geralt M, Serrano P, Mohanty B, Horst R, Pedrini B, Elsliger MA, Wilson IA, Wüthrich K (2010) NMR structure of the protein NP_247299.1: comparison with the crystal structure. Acta Cryst F 66:1367–1380CrossRefGoogle Scholar
  15. Keller R (2004) Computer-aided resonance assignment. Cantina.
  16. Koradi R, Billeter M, Wüthrich K (1996) MOLMOL: a program for display and analysis of macromolecular structures. J Mol Graph 14(51–55):29–32Google Scholar
  17. Koradi R, Billeter M, Güntert P (2000) Point-centered domain decomposition for parallel molecular dynamics simulation. Comp Phys Commun 124:139–147ADSCrossRefzbMATHGoogle Scholar
  18. Laskowski RA, Macarthur MW, Moss DS, Thornton JM (1993) PROCHECK: a program to check the stereochemical quality of protein structures. J Appl Cryst 26:283–291CrossRefGoogle Scholar
  19. Lloyd NR, Wuttke DS (2014) Less is more: structures of difficult targets with minimal constraints. Structure 22:1223–1224CrossRefGoogle Scholar
  20. Luginbühl P, Güntert P, Billeter M, Wüthrich K (1996) The new program OPAL for molecular dynamics simulations and energy refinements of biological macromolecules. J Biomol NMR 8:136–146CrossRefGoogle Scholar
  21. Mohanty B, Serrano P, Pedrini B, Jaudzems K, Geralt M, Horst R, Herrmann T, Elsliger MA, Wilson IA, Wüthrich K (2010) Comparison of NMR and crystal structures of the proteins TM1112 and TM1367. Acta Cryst F 66:1381–1392CrossRefGoogle Scholar
  22. Mohanty B, Serrano P, Geralt M, Wüthrich K (2014) NMR structure determination of the protein NP_344798.1 as the first representative of the Pfam PF06042. J Biomol NMR (in press)Google Scholar
  23. Pedrini B, Serrano P, Mohanty B, Geralt M, Wüthrich K (2013) NMR-profiles of protein solutions. Biopolymers 99:825–831CrossRefGoogle Scholar
  24. Serrano P, Pedrini B, Mohanty B, Geralt M, Herrmann T, Wüthrich K (2010) Comparison of NMR and crystal structures highlights conformational isomerism in protein active sites. Acta Cryst F 66:1393–1406CrossRefGoogle Scholar
  25. Serrano P, Pedrini B, Geralt M, Jaudzems K, Mohanty B, Horst R, Herrmann T, Wüthrich K (2012) The J-UNIO protocol for automated protein structure determination by NMR in solution. J Biomol NMR 53:341–354CrossRefGoogle Scholar
  26. Serrano P, Geralt M, Mohanty B, Wüthrich K (2014) NMR structures of the α-proteobacterial ATPase-regulating ζ-subunits. J Mol Biol 15:2547–2553CrossRefGoogle Scholar
  27. Sgourakis NG, Natajaran K, Ying J, Vögeli B, Boyd LF, Margulies DH, Bax A (2014) The structure of mouse cytomegalovirus m04 protein obtained from sparse NMR data reveals a conserved fold of the m02–m06 viral immune modulator family. Structure 22:1263–1273CrossRefGoogle Scholar
  28. Strange RW, Antonyuk SV, Ellis MJ, Bessho Y, Kuramitsu S, Shinkai A, Yokoyama S, Hasnain SS (2009) Structure of a putative beta-phosphoglucomutase (TM1254) from Thermotoga maritima. Acta Cryst F 65:1218–1221CrossRefGoogle Scholar
  29. Volk J, Herrmann T, Wüthrich K (2008) Automated sequence-specific protein NMR assignment using the memetic algorithm MATCH. J Biomol NMR 41:127–138CrossRefGoogle Scholar
  30. Wahab AT, Serrano P, Geralt M, Wüthrich K (2011) NMR structure of the Bordetella Bronchiseptica protein NP_888769.1 establishes a new phage-related protein family PF13554. Prot Sci 20:1137–1144CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2014

Authors and Affiliations

  • Kristaps Jaudzems
    • 1
    • 2
    • 5
  • Bill Pedrini
    • 1
    • 2
    • 4
    • 6
  • Michael Geralt
    • 1
    • 2
  • Pedro Serrano
    • 1
    • 2
  • Kurt Wüthrich
    • 1
    • 2
    • 3
    • 4
    Email author
  1. 1.Department of Integrative Structural and Computational BiologyThe Scripps Research InstituteLa JollaUSA
  2. 2.Joint Center for Structural GenomicsLa JollaUSA
  3. 3.The Skaggs Institute for Chemical BiologyThe Scripps Research InstituteLa JollaUSA
  4. 4.Institute of Molecular Biology and BiophysicsETH ZürichZurichSwitzerland
  5. 5.Latvian Institute of Organic SynthesisRigaLatvia
  6. 6.SwissFEL ProjectPaul Scherrer Institute (PSI)VilligenSwitzerland

Personalised recommendations