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The formation of a native-like structure containing eight conserved hydrophobic residues is rate limiting in two-state protein folding of ACBP

Nature Structural Biology volume 6, pages 594–601 (1999)Cite this article

Abstract

The acyl-coenzyme A-binding proteins (ACBPs) contain 26 highly conserved sequence positions. The majority of these have been mutated in the bovine protein, and their influence on the rate of two-state folding and unfolding has been measured. The results identify eight sequence positions, out of 24 probed, that are critical for fast productive folding. The residues are all hydrophobic and located in the interface between the N- and C-terminal helices. The results suggest that one specific site dominated by conserved hydrophobic residues forms the structure of the productive rate-determining folding step and that a sequential framework model can describe the protein folding reaction.

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Figure 1: a, Three-dimensional structure of bovine ACBP (PDB accession code 2ABD).
Figure 2: a, Equilibrium unfolding of ACBP variants at 278 K, 0.02 M sodium acetate, pH 5.3.
Figure 3: Structure of ACBP emphasizing the eight residues that are critical for formation of RLNLS.
Figure 4: Correlation between ΔΔGD-N and ΔΔGD-RLNLS showing two groups of variants.
Figure 5: The sequential framework model for ACBP folding.

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Acknowledgements

K. Hansen, J. Stenvang Jepsen, K. Poulsen, S. Radmehr and P.V. Sørensen are acknowledged for production of some ACBP variants. E. Knudsen, J. Andersen and P. Skovgaard are acknowledged for skilled technical assistance, and C. Rischel is thanked for providing the program CANOO. We acknowledge the advice to make additional mutations from the referees of this paper. This publication is a contribution from the Danish Protein Engineering Research Center and is also supported by the Danish Natural Research Councils.

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Author notes

  1. Birthe B. Kragelund & Flemming M. Poulsen

    Present address: Institute of Molecular Biology, University of Copenhagen, Øster Farimagsgade 2A, DK-1353, Copenhagen, Denmark

  2. Peter Osmark

    Present address: Department of Biochemistry, University of Montreal, 6128, Quebec, Canada

Authors and Affiliations

  1. Carlsberg Laboratorium, Gamle Carlsberg Vej 10, DK-2500 Valby, Copenhagen, Denmark

    Birthe B. Kragelund, Peter Osmark, Jacob Schiødt & Flemming M. Poulsen

  2. Institute of Biochemistry, Odense University, Campusvej, 55 DK-5230, Odense M, Denmark

    Thomas B. Neergaard & Jens Knudsen

  3. Department of Molecular Biology, Odense University, Campusvej, 55 DK-5230, Odense M, Denmark

    Karsten Kristiansen

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Correspondence to Flemming M. Poulsen.

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Kragelund, B., Osmark, P., Neergaard, T. et al. The formation of a native-like structure containing eight conserved hydrophobic residues is rate limiting in two-state protein folding of ACBP. Nat Struct Mol Biol 6, 594–601 (1999). https://doi.org/10.1038/9384

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