The structure of the transition state for the association of two fragments of the barley chymotrypsin inhibitor 2 to generate native-like protein: implications for mechanisms of protein folding

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 91; no. 23; pp. 10943 - 10946
• Main Authors: de Prat Gay, G, Ruiz-Sanz, J, Davis, B, Fersht, A R
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences of the United States of America 08.11.1994; National Acad Sciences; National Academy of Sciences
• Subjects: Amino Acid Sequence; Association reactions; Barley; Biochemistry; chymotrypsin; Chymotrypsin - antagonists & inhibitors; chymotrypsine; Circular Dichroism; Dichroism; enzyme inhibitors; Free energy; Hordeum - enzymology; hordeum vulgare; inhibidores de enzimas; inhibiteur d' enzyme; Kinetics; Magnetic Resonance Spectroscopy; Molecular Sequence Data; Mutagenesis, Site-Directed; Mutation; Peptides; Plant Proteins - chemistry; Protein engineering; Protein Folding; Protein Structure, Secondary; Protein Structure, Tertiary; proteinas; proteine; Proteins; quimotripsina; Serine Proteinase Inhibitors - chemistry; Spectroscopy; Structure-Activity Relationship; Thermodynamics
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.91.23.10943

Possible early events in protein folding may be studied by dissecting proteins into complementary fragments. Two fragments of chymotrypsin inhibitor 2 [CI2-(20-59) and CI2-(60-83)] associate to form a native-like structure in a second-order reaction that combines collision and rearrangement. The transition state of the reaction, analyzed by the protein engineering method on 17 mutants, is remarkably similar to that for the folding of the intact protein--a structure that resembles an expanded version of the folded structure with most interactions significantly weakened. The exception is that the N-terminal region of the single a-helix (the N-capping box) is completely formed in the transition state for association of the fragments, whereas it is reasonably well formed for the intact protein. Preliminary evidence on the structures of the individual fragments indicates that both are mainly nonnative, lacking native secondary structure and having regions of nonnative buried hydrophobic clusters. The association reaction does not result from the collision of a subpopulation of two fully native-like fragments but involves a considerable rearrangement of structure.