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Conservation of folding pathways in evolutionarily distant globin sequences

Nature Structural Biology volume 7, pages 679–686 (2000)Cite this article

Abstract

To test the hypothesis that the folding pathways of evolutionarily related proteins with similar three-dimensional structures but widely different sequences should be similar, the folding pathway of apoleghemoglobin has been characterized using stopped-flow circular dichroism, heteronuclear NMR pulse labeling techniques and mass spectrometry. The pathway of folding was found to differ significantly from that of a protein of the same family, apomyoglobin, although both proteins appear to fold through helical burst phase intermediates. For leghemoglobin, the burst phase intermediate exhibits stable helical structure in the G and H helices, together with a small region in the center of the E helix. The A and B helices are not stabilized until later stages of the folding process. The structure of the burst phase folding intermediate thus differs from that of apomyoglobin, in which stable helical structure is formed in the A, B, G and H helix regions.

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Figure 1: Comparison of the sequences of Mb and Lb.
Figure 2: Comparison of the pH and urea dependent behaviors of apoLb and apoMb.
Figure 3: Kinetics of apoLb refolding from urea.
Figure 4: 15N-1H HSQC spectrum of LbCO after reconstitution.
Figure 5: Summary of the quench-flow results for Lb.
Figure 6: Effect of labeling pulse strength and duration.
Figure 7: Mass spectrometric analysis of the folding pathway of apoLb.
Figure 8: Stereo view of the backbone structure of soybean Lb a17.

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Acknowledgements

We thank J. Chung for assistance with the NMR experiments, D. Eliezer, J. Yao, P. Jennings, C. Garcia, V. Tsui and S. Cavagnero for helpful discussions, and L. Tennant for expert technical assistance. This work was supported by grants from the National Institutes of Health. S.P. was supported by a fellowship from the Deutsches Forschungsgemeinschaft.

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Authors and Affiliations

  1. Department of Molecular Biology and Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, 92037, California, USA

    Chiaki Nishimura, H. Jane Dyson & Peter E. Wright

  2. Institut für Physikalische Chemie, Karl-Franzens Universität Graz, Universitätsplatz 1, A-8010, Austria

    Stefan Prytulla

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  1. Chiaki Nishimura
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Correspondence to H. Jane Dyson or Peter E. Wright.

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Nishimura, C., Prytulla, S., Jane Dyson, H. et al. Conservation of folding pathways in evolutionarily distant globin sequences. Nat Struct Mol Biol 7, 679–686 (2000). https://doi.org/10.1038/77985

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