Solution NMR evidence for a cis Tyr-Ala peptide group in the structure of [Pro93Ala] bovine pancreatic ribonuclease A

Protein Sci. 2000 Feb;9(2):421-6. doi: 10.1110/ps.9.2.421.


Proline peptide group isomerization can result in kinetic barriers in protein folding. In particular, the cis proline peptide conformation at Tyr92-Pro93 of bovine pancreatic ribonuclease A (RNase A) has been proposed to be crucial for chain folding initiation. Mutation of this proline-93 to alanine results in an RNase A molecule, P93A, that exhibits unfolding/refolding kinetics consistent with a cis Tyr92-Ala93 peptide group conformation in the folded structure (Dodge RW, Scheraga HA, 1996, Biochemistry 35:1548-1559). Here, we describe the analysis of backbone proton resonance assignments for P93A together with nuclear Overhauser effect data that provide spectroscopic evidence for a type VI beta-bend conformation with a cis Tyr92-Ala93 peptide group in the folded structure. This is in contrast to the reported X-ray crystal structure of [Pro93Gly]-RNase A (Schultz LW, Hargraves SR, Klink TA, Raines RT, 1998, Protein Sci 7:1620-1625), in which Tyr92-Gly93 forms a type-II beta-bend with a trans peptide group conformation. While a glycine residue at position 93 accommodates a type-II bend (with a positive value of phi93), RNase A molecules with either proline or alanine residues at this position appear to require a cis peptide group with a type-VI beta-bend for proper folding. These results support the view that a cis Pro93 conformation is crucial for proper folding of wild-type RNase A.

Publication types

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

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Cattle
  • Magnetic Resonance Spectroscopy
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Pancreas / enzymology
  • Proline / chemistry
  • Protein Conformation
  • Protein Folding
  • Protein Structure, Secondary
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Ribonuclease, Pancreatic / chemistry*
  • Ribonuclease, Pancreatic / genetics
  • Solutions


  • Recombinant Proteins
  • Solutions
  • Proline
  • Ribonuclease, Pancreatic