Protein folding and stability using denaturants.
Measurements of protein folding and thermodynamic stability provide insight into the forces and energetics that determine structure, and can inform on protein domain organization, interdomain interactions, and effects of mutations on structure. This chapter describes methods, theory, and data analysis for the most accessible means to determine the thermodynamics of protein folding: chemical denaturation. Topics include overall features of the folding reaction, advances in instrumentation, optimization of reagent purity, mechanistic models for analysis, and statistical and structural interpretation of fitted thermodynamic parameters. Examples in which stability measurements have provided insight into structure and function will be taken from studies in the author's laboratory on the Notch signaling pathway. It is hoped that this chapter will enable molecular, cell, and structural biologists to make precise measurements of protein stability, and will also provide a strong foundation for biophysics students who wish to undertake experimental studies of protein folding.
Street
Timothy O
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T. C. Jenkins Department of Biophysics, The Johns Hopkins University, Baltimore, Maryland 21218, USA.
Courtemanche
Naomi
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Barrick
Doug
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D000818:Animals; D004330:Drosophila; D008954:Models, Biological; D011489:Protein Denaturation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D051880:Receptors, Notch / Q000737:chemistry; D012996:Solutions; D013816:Thermodynamics; D017186:Titrimetry; D014508:Urea
D051880:Receptors, Notch; D012996:Solutions; D014508:Urea