Characterizing the residue level folding of the intrinsically unstructured IA3

Biochemistry. 2006 Nov 14;45(45):13585-96. doi: 10.1021/bi061358w.


Residue level analysis of the folding of simple proteins may hold the key to understanding folding pathways and aid in structure prediction. IA(3), the endogenous inhibitor of yeast aspartic proteinase A (YPrA), is an unstructured protein in solution. Comparison of the 2D (15)N-HSQC spectra of IA(3) in water and in 23% 2,2,2-trifluoroethanol (TFE) shows that the individual residue cross peaks of IA(3) become more dispersed in the presence of TFE, indicating that the protein undergoes an unstructured to structured transition in the presence of TFE. This transition can be monitored by the movements of the cross peaks. Following the individual cross peaks, however, is complicated and does not establish whether a single transition occurs globally in the sequence. In this equilibrium study, we apply singular value decomposition (SVD) to elucidate both the main features of the TFE-driven transition and the residue-level deviations from the average behavior. This analysis has yielded a two-state folding description as well as specifics of NMR frequency shifts of individual residues, indicating that the N-terminus of IA(3) has a higher helical propensity than the C-terminus. Additionally, we discuss possible mechanisms for observed deviations from a two-state folding transition. When combined with a traditional biochemical understanding of interactions between individual residues, this approach leads to a better understanding of protein folding.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Aspartic Acid Endopeptidases / antagonists & inhibitors
  • Deoxyribonucleases, Type II Site-Specific / chemistry*
  • Deoxyribonucleases, Type II Site-Specific / drug effects
  • Nuclear Magnetic Resonance, Biomolecular
  • Protein Folding*
  • Saccharomyces cerevisiae Proteins / chemistry*
  • Saccharomyces cerevisiae Proteins / drug effects
  • Thermodynamics
  • Trifluoroethanol / chemistry
  • Trifluoroethanol / pharmacology


  • Saccharomyces cerevisiae Proteins
  • Trifluoroethanol
  • AI3 protein, S cerevisiae
  • Deoxyribonucleases, Type II Site-Specific
  • Aspartic Acid Endopeptidases