Folding propensities of synthetic peptide fragments covering the entire sequence of phage 434 Cro protein

The phage 434 Cro protein, the N-terminal domain of its repressor (R1-69) a nd that of phage lambda (lambda(6-85)) constitute a group of small, monomer ic, single-domain folding units consisting of five helices with striking st ructural similarity. The intrinsic helix stabilities in lambda(6-85) have b een correlated to its rapid folding behavior, and a residual hydrophobic cl uster 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 com parison with R1-69 and lambda(6-85), we examined the conformational behavio r of five peptides covering the entire 434 Cro sequence in water, 40% (by v olume) TFE/water, and 7 M urea solutions using CD and NMR. Each peptide cor responds to a helix and adjacent residues as identified in the native 434 C ro NMR and crystal structures. All are soluble and monomeric in the solutio n conditions examined except for the peptide corresponding to the 434 Cro h elix 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 t he 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 fold ing propensities of the five helices could account for the observed stabili ty and folding behavior of 434 Cro and is, at least qualitatively, in accor d with the results of the recently described diffusion-collision model inco rporating intrinsic helix propensities.