Computer-based redesign of a protein folding pathway

A fundamental test of our current understanding of protein folding is to rationally redesign protein folding pathways. We use a computer-based design strategy to switch the folding pathway of protein G, which normally involves formation of the second, but not the first, β-turn at the rate limiting step in folding. Backbone conformations and amino acid sequences that maximize the interaction density in the first β-hairpin were identified, and two variants containing 11 amino acid replacements were found to be ∼4 kcal mol−1 more stable than wild type protein G. Kinetic studies show that the redesigned proteins fold ∼100× faster than wild type protein and that the first β-turn is formed and the second disrupted at the rate limiting step in folding.

[1]  S. Honda,et al.  Role of side-chains in the cooperative beta-hairpin folding of the short C-terminal fragment derived from streptococcal protein G. , 2000, Biochemistry.

[2]  L. Serrano,et al.  A short linear peptide that folds into a native stable β-hairpin in aqueous solution , 1994, Nature Structural Biology.

[3]  B. L. Sibanda,et al.  β-Hairpin families in globular proteins , 1985, Nature.

[4]  G. Rose,et al.  Is protein folding hierarchic? II. Folding intermediates and transition states. , 1999, Trends in biochemical sciences.

[5]  Lorna J. Smith,et al.  Understanding protein folding via free-energy surfaces from theory and experiment. , 2000, Trends in biochemical sciences.

[6]  A. Finkelstein,et al.  A theoretical search for folding/unfolding nuclei in three-dimensional protein structures. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[7]  A. Fersht,et al.  The folding of an enzyme. I. Theory of protein engineering analysis of stability and pathway of protein folding. , 1992, Journal of molecular biology.

[8]  J. Onuchic,et al.  The energy landscape theory of protein folding: insights into folding mechanisms and scenarios. , 2000, Advances in protein chemistry.

[9]  V. Muñoz,et al.  A simple model for calculating the kinetics of protein folding from three-dimensional structures. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[10]  A. Gronenborn,et al.  A novel, highly stable fold of the immunoglobulin binding domain of streptococcal protein G. , 1993, Science.

[11]  D. Baker,et al.  Critical role of β-hairpin formation in protein G folding , 2000, Nature Structural Biology.

[12]  D. Baker,et al.  Native protein sequences are close to optimal for their structures. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[13]  F. Young.  Biochemistry , 1955, The Indian Medical Gazette.

[14]  D. Baker,et al.  Contact order, transition state placement and the refolding rates of single domain proteins. , 1998, Journal of molecular biology.

[15]  D Baker,et al.  A breakdown of symmetry in the folding transition state of protein L. , 2000, Journal of molecular biology.

[16]  Terrence G. Oas,et al.  The energy landscape of a fast-folding protein mapped by Ala→Gly Substitutions , 1997, Nature Structural Biology.

[17]  Stephen L. Mayo,et al.  Design, structure and stability of a hyperthermophilic protein variant , 1998, Nature Structural Biology.

[18]  D. Baker,et al.  A surprising simplicity to protein folding , 2000, Nature.

[19]  A. Fersht.  Structure and mechanism in protein science , 1998 .

[20]  G L Gilliland,et al.  Two crystal structures of the B1 immunoglobulin-binding domain of streptococcal protein G and comparison with NMR. , 1994, Biochemistry.

[21]  Shoji Takada,et al.  Variational Theory for Site Resolved Protein Folding Free Energy Surfaces , 1998, cond-mat/9805366.