Va. Jaravine et al., Observation of the closing of individual hydrogen bonds during TFE-inducedhelix formation in a peptide, PROTEIN SCI, 10(5), 2001, pp. 943-950
Helix formation of an S-peptide analog, comprising the first 20 residues of
Ribonuclease A and two additional N-terminal residues, was studied by meas
uring hydrogen bond (H-bond) (h3)J(NC') scalar couplings as a function of 2
,2,2-trifluoroethanol (TFE) concentration. The (h3)J(NC') couplings give di
rect evidence for the closing of individual backbone N-H . .O=C H-bonds dur
ing the TFE-induced formation of secondary structure. Whereas no (h3)J(NC')
correlations could be detected without TFE, alpha -helical (i,i +4) H-bond
correlations were observed for the amides of residues A5 to M15 in the pre
sence of TFE. The analysis of individual coupling constants indicates that
a-helix formation starts at the center of the S-peptide around residue Ell
and proceeds gradually from there to both peptide ends as the TFE concentra
tion is increased. At 60% to 90% TFE, well-formed a-helical H-bonds were ob
served for the amides hydrogens of residues K9 to Q13, whereas H-bonds of r
esidues T5 to A8, H14, and M15 are affected by fraying. No intramolecular b
ackbone H-bonds are present at and beyond the putative helix stop signal D1
6. As the (h3)J(NC') constants represent ensemble averages and the dependen
ce of (h3)J(NC') on H-bond lengths is very steep, the size of the individua
l (h3)J(NC') coupling constants can be used as a measure for the population
of a closed H-bond. These individual populations sire in agreement with re
sults derived from the Lifson-Roig theory for coil-to-helix transitions. Th
e present work shows that the closing of individual H-bonds during TFE-indu
ced helix formation can be monitored by changes in the size of H-bond scala
r couplings.