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Effect of disulfide bridge formation on the NMR spectrum of a protein: Studies on oxidized and reduced Escherichia coli thioredoxin

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Summary

As a prelude to complete structure calculations of both the oxidized and reduced forms of Escherchia coli thioredoxin (Mr 11 700), we have analyzed the NMR data obtained for the two proteins under identical conditions. The complete aliphatic 13C assignments for both oxidized and reduced thioredoxin are reported. Correlations previously noted between 13C chemical shifts and secondary structure are confirmed in this work, and significant differences are observed in the Cβ and Cγ shifts between cis- and trans-proline, consistent with previous work that identifies this as a simple and unambiguous method of identifying cis-proline residues in proteins. Reduction of the disulfide bond in the active-site Cys32-Gly-Pro-Cys35 sequence causes changes in the 1H, 15N and 13C chemical shifts of residues close to the active site, some of them quite far distant in the amino acid sequence. Coupling constants, both backbone and side chain, show some differences between the two proteins, and the NOE connectivities and chemical shifts are consistent with small changes in the positions of several side chains, including the two tryptophan rings (Trp28 and Trp31). These results show that, consistent with the biochemical behavior of thioredoxin, there are minimal differences in backbone configuration between the oxidized and reduced forms of the protein.

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  1. K. Chandrasekhar

    Present address: Harvard Medical School, 02115, Boston, MA, USA

Authors and Affiliations

  1. Department of Molecular Biology, The Scripps Research Institute, 10666 North Torrey Pines Road, 92037, La Jolla, CA, USA

    K. Chandrasekhar, A. Patricia Campbell, Mei-Fen Jeng & H. Jane Dyson

  2. Karolinska Institute, S-10401, Stockholm, Sweden

    Arne Holmgren

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Chandrasekhar, K., Campbell, A.P., Jeng, MF. et al. Effect of disulfide bridge formation on the NMR spectrum of a protein: Studies on oxidized and reduced Escherichia coli thioredoxin. J Biomol NMR 4, 411–432 (1994). https://doi.org/10.1007/BF00179349

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  • DOI: https://doi.org/10.1007/BF00179349

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