The phage 434 Cro protein, the N-terminal domain
of its repressor (R1–69) and that of phage λ
(λ6–85) constitute a group of small,
monomeric, single-domain folding units consisting of five
helices with striking structural similarity. The intrinsic
helix stabilities in λ6–85 have been
correlated to its rapid folding behavior, and a residual
hydrophobic cluster found in R1–69 in 7 M urea has
been proposed as a folding initiation site. To understand
the early events in the folding of 434 Cro, and for comparison
with R1–69 and λ6–85, we examined
the conformational behavior of five peptides covering the
entire 434 Cro sequence in water, 40% (by volume) TFE/water,
and 7 M urea solutions using CD and NMR. Each peptide corresponds
to a helix and adjacent residues as identified in the native
434 Cro NMR and crystal structures. All are soluble and
monomeric in the solution conditions examined except for
the peptide corresponding to the 434 Cro helix 4, which
has low water solubility. Helix formation is observed for
the 434 Cro helix 1 and helix 2 peptides in water, for
all the peptides in 40% TFE and for none in 7 M urea. NMR
data indicate that the helix limits in the peptides are
similar to those in the native protein helices. The number
of side-chain NOEs in water and TFE correlates with the
helix content, and essentially none are observed in 7 M
urea for any peptide, except that for helix 5, where a
hydrophobic cluster may be present. The low intrinsic folding
propensities of the five helices could account for the
observed stability and folding behavior of 434 Cro and
is, at least qualitatively, in accord with the results
of the recently described diffusion-collision model incorporating
intrinsic helix propensities.
