Populating partially unfolded forms by hydrogen exchange-directed protein engineering.

Jiro Takei; Wuhong Pei; Diep Vu; Yawen Bai

doi:10.1021/bi026491c

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Populating partially unfolded forms by hydrogen exchange-directed protein engineering.

Jiro Takei·Wuhong Pei·Diep Vu·Yawen Bai

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Abstract

The native-state hydrogen exchange of a redesigned apocytochrome b(562) suggests that at least two partially unfolded forms (PUFs) exist for this four-helix bundle protein under native conditions. The more stable PUF has the N-terminal helix unfolded. To verify the conclusion further and obtain more detailed structural information about this PUF, five hydrophobic core residues in the N-terminal helix were mutated to Gly and Asp to destabilize the native state selectively and populate the PUF for structural studies. The secondary structure and the backbone dynamics of this mutant were characterized using multidimensional NMR. Consistent with the prediction, the N-terminal region of the mutant was found to be unfolded while other parts of the proteins remained folded. These results suggest that native-state hydrogen exchange-directed protein engineering can be a useful approach to populating partially unfolded forms for detailed structural studies.

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Amyloid Apoproteins Circular Dichroism Cytochrome b Group Glycine Hydrogen Mutagenesis, Site-Directed Nuclear Magnetic Resonance, Biomolecular Protein Conformation Protein Engineering Protein Folding Protein Structure, Secondary Thermodynamics Journal Article

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Jiro Takei, Wuhong Pei, Diep Vu, Yawen Bai. 2002-10-15. Populating partially unfolded forms by hydrogen exchange-directed protein engineering.. https://doi.org/10.1021/bi026491c

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