Observation of the closing of individual hydrogen bonds during TFE–induced helix formation in a peptide

• Published in: Protein science 2001-05, Vol.10 (5), p.943-950
• Main Authors: Jaravine, Victor A., Alexandrescu, Andrei T., Grzesiek, Stephan
• Format: Article
• Language: English
• Subjects: Amides - chemistry; Amides - metabolism; Amino Acid Sequence; Dose-Response Relationship, Drug; Enzyme Stability - drug effects; Hydrogen Bonding - drug effects; H‐bond, hydrogen bond; J‐coupling; Magnetic Resonance Spectroscopy; Molecular Sequence Data; NMR; Peptide Fragments - chemical synthesis; Peptide Fragments - chemistry; Peptide Fragments - metabolism; Protein Denaturation - drug effects; Protein Folding; Protein Structure, Secondary - drug effects; Ribonuclease, Pancreatic - chemistry; RnaseA; RNaseA, Ribonuclease A; scalar coupling; S‐peptide; S‐peptide, N‐terminal fragment of Ribonuclease A; TFE, 2,2,2‐trifluoroethanol; Thermodynamics; Trifluoroethanol - pharmacology
• ISSN: 0961-8368; 1469-896X
• DOI: 10.1110/ps.48501

Helix formation of an S‐peptide analog, comprising the first 20 residues of Ribonuclease A and two additional N‐terminal residues, was studied by measuring hydrogen bond (H‐bond) h3JNC′ scalar couplings as a function of 2,2,2‐trifluoroethanol (TFE) concentration. The h3JNC′ couplings give direct evidence for the closing of individual backbone N‐H•••O = C H‐bonds during the TFE‐induced formation of secondary structure. Whereas no h3JNC′ correlations could be detected without TFE, α‐helical (i,i +4) H‐bond correlations were observed for the amides of residues A5 to M15 in the presence of TFE. The analysis of individual coupling constants indicates that α‐helix formation starts at the center of the S‐peptide around residue E11 and proceeds gradually from there to both peptide ends as the TFE concentration is increased. At 60% to 90% TFE, well‐formed α‐helical H‐bonds were observed for the amides hydrogens of residues K9 to Q13, whereas H‐bonds of residues T5 to A8, H14, and M15 are affected by fraying. No intramolecular backbone H‐bonds are present at and beyond the putative helix stop signal D16. As the h3JNC′ constants represent ensemble averages and the dependence of h3JNC′ on H‐bond lengths is very steep, the size of the individual h3JNC′ coupling constants can be used as a measure for the population of a closed H‐bond. These individual populations are in agreement with results derived from the Lifson‐Roig theory for coil‐to‐helix transitions. The present work shows that the closing of individual H‐bonds during TFE‐induced helix formation can be monitored by changes in the size of H‐bond scalar couplings.