Skip to main content
SpringerLink
Log in

Measurement of 15N-1H coupling constants in uniformly 15N-labeled proteins: Application to the photoactive yellow protein

  • Published:
Journal of Biomolecular NMR Aims and scope Submit manuscript

Abstract

A modified HNHB experiment is presented that allows thedetermination of J(NH) coupling constants directly from the ratio ofcross-peak to diagonal-peak intensities. The experiment was applied to thephotoactive yellow protein (PYP) and yielded the magnitude of 1173J(NHβ) coupling constants. In addition, 293J(NHα(i−1)) coupling constantscould be measured, providing information about the backbone angle ψ.These data, in conjunction with the magnitudes of the3J(HNHα) coupling constantsobtained from the HNHA spectrum, effectively discriminate the twopossibilities for the stereospecific assignment of theHα resonances in glycine residues. For all eight glycineresidues in PYP that were not subject to conformational averaging and hadnon-degenerate Hα resonance frequencies, the J-couplingdata, together with limited NOE data, yielded the stereospecific assignmentof the Hα resonances for these residues. In addition,reliable and precise φ,ψ dihedral constraints were also derived forthese residues from the J-coupling data.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  • Archer, S.J., Ikura, M., Torchia, D.A. and Bax, A. (1991) J. Magn. Reson., 95, 636–641.

    Google Scholar 

  • Bax, A., Vuister, G.W., Grzesiek, S., Delaglio, F., Wang, A.C., Tschudin, R. and Zhu, G. (1994) Methods Enzymol., 239, 79–105.

    Google Scholar 

  • Bystrov, V.F. (1976) Prog. NMR Spectrosc., 10, 41–81.

    Google Scholar 

  • Delaglio, F., Grzesiek, S., Vuister, G.W., Zhu, G., Pfeifer, J. and Bax, A. (1995) J. Biomol. NMR, 6, 277–293.

    Google Scholar 

  • Emerson, S.D. and Montelione, G.T. (1992) J. Magn. Reson., 99, 413–420.

    Google Scholar 

  • Gemmecker, G. and Fesik, S.W. (1991) J. Magn. Reson., 95, 208–213.

    Google Scholar 

  • Griesinger, C., Sørensen, O.W. and Ernst, R.R. (1986) J. Chem. Phys., 85, 6837–6843.

    Google Scholar 

  • Griesinger, C. and Eggenberger, U. (1992) J. Magn. Reson., 97, 426–434.

    Google Scholar 

  • Karimi-Nejad, Y., Schmidt, J.M., Rüterjans, H., Schwalbe, H. and Griesinger, C. (1994) Biochemistry, 33, 5481–5492.

    Google Scholar 

  • Kleywegt, G.J., Vuister, G.W., Padilla, A., Knechtel, R.M., Boelens, R. and Kaptein, R. (1993) J. Magn. Reson., 102, 166–176.

    Google Scholar 

  • Madsen, J.C., Sørensen, O.W., Sørensen, P. and Poulsen, F.M. (1993) J. Biomol. NMR, 3, 239–244.

    Google Scholar 

  • Montelione, G.T., Winkler, M.E., Rauenbuehler, P. and Wagner, G. (1989) J. Magn. Reson., 82, 198–204.

    Google Scholar 

  • Pardi, A., Billeter, M. and Wüthrich, K. (1984) J. Mol. Biol., 180, 741–751.

    Google Scholar 

  • Seip, S., Balbach, J. and Kessler, H. (1994) J. Magn. Reson., B104, 172–179.

    Google Scholar 

  • Shaka, A.J., Keeler, J., Frenkiel, T. and Freeman, R. (1983) J. Magn. Reson., 52, 335–338.

    Google Scholar 

  • Sklenar, V., Piotto, M., Leppik, R. and Saudek, V. (1993) J. Magn. Reson., A102, 241–245.

    Google Scholar 

  • Sørensen, O.W., Eich, G.W., Levitt, M.H., Bodenhausen, G. and Ernst, R.R. (1983) Prog. NMR Spectrosc., 16, 163–192.

    Google Scholar 

  • Tjandra, N., Grzesiek, S. and Bax, A. (1996) J. Am. Chem. Soc., 118, 6264–6272.

    Google Scholar 

  • Tolman, J.R., Flanagan, J.M., Kennedy, M.A. and Prestegard, J.H. (1995) Proc. Natl. Acad. Sci. USA, 92, 9279–9283.

    Google Scholar 

  • Vuister, G.W. and Bax, A. (1993a) J. Am. Chem. Soc., 115, 7772–7777.

    Google Scholar 

  • Vuister, G.W. and Bax, A. (1993b) J. Magn. Reson., B102, 228–231.

    Google Scholar 

  • Vuister, G.W., Yamazaki, T., Torchia, D.A. and Bax, A. (1993) J. Biomol. NMR, 3, 297–306.

    Google Scholar 

  • Wang, A.C. and Bax, A. (1995) J. Am. Chem. Soc., 117, 1810–1813.

    Google Scholar 

  • Wang, A.C. and Bax, A. (1996) J. Am. Chem. Soc., 118, 2483–2494.

    Google Scholar 

  • Weisemann, R., Rüterjans, H., Schwalbe, H., Schleucher, J., Bermel, W. and Griesinger, C. (1994) J. Biomol. NMR, 4, 231–240.

    Google Scholar 

  • Wider, G., Neri, D., Otting, G. and Wüthrich, K. (1989) J. Magn. Reson., 85, 426–431.

    Google Scholar 

  • Xu, R.X., Olejniczak, E.T. and Fesik, S.W. (1992) FEBS Lett., 305, 137–143.

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of NMR Spectroscopy, Bijvoet Center for Biomolecular Research, Utrecht University, Padualaan 8, 3584 CH, Utrecht, The Netherlands

    Petra Düx, Brian Whitehead, Rolf Boelens, Robert Kaptein & Geerten W. Vuister

Authors
  1. Petra Düx
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Brian Whitehead
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Rolf Boelens
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Robert Kaptein
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Geerten W. Vuister
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Düx, P., Whitehead, B., Boelens, R. et al. Measurement of 15N-1H coupling constants in uniformly 15N-labeled proteins: Application to the photoactive yellow protein. J Biomol NMR 10, 301–306 (1997). https://doi.org/10.1023/A:1018393225804

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/A:1018393225804

Search

Navigation