Perturbation of peptide conformations induced in anisotropic environments

Abstract

Reversed-phase high performance liquid chromatography (RP HPLC) has been found to be a convenient and powerful tool for the study of the secondary structure of peptides. Here, the ability of proline to perturb the secondary structures of peptides induced at aqueous-lipid interfaces and the induced conformation of polyproline peptides were investigated by means of RP HPLC. For these studies, four different complete sets of substitution analogues of model peptides expected to have specific induced conformations were used. In the first two studies, a single lysine was "walked" through two 18-residue polyproline sequences (one N-acetylated, the other not). In the remaining two studies, a proline was "walked" through two different sequences that had been found earlier to be induced into an alpha-helical conformation during RP HPLC (an 18-residue polyalanine sequence and the amphipathic 14-residue sequence Ac-LLKLLKKLLKKLKK-NH2). Sixty-eight individual analogues were synthesized for this study and the effect of the respective substitutions on retention times was determined. The results are consistent with the concept that, upon interaction with the C-18 of the stationary phase during RP HPLC, polyproline is induced into a type II helical conformation, polyalanine into an alpha-helical conformation, and Ac-LLKLLKKLLKKLKK-NH2 into an amphipathic alpha-helical array. In an extension of this study, the antimicrobial activities of Ac-LLKLLKKLLKKLKK-NH2 and its 18 proline substitution analogues were found to be inversely correlated with their RP HPLC retention times.

Melectin: A Novel Antimicrobial Peptide from the Venom of the Cleptoparasitic Bee Melecta albifrons

“…[34] Although RP-HPLC separates peptides mainly by hydrophobic interactions, the induced peptide secondary structures also influence their retention times. [34] The substitution of Lys for Pro in MEP-1 reduced the hydrophobicity of the peptide in a way that theoretically should shorten its retention time measured on the C-18 RP-HLC column.…”

“…[34] Although RP-HPLC separates peptides mainly by hydrophobic interactions, the induced peptide secondary structures also influence their retention times. [34] The substitution of Lys for Pro in MEP-1 reduced the hydrophobicity of the peptide in a way that theoretically should shorten its retention time measured on the C-18 RP-HLC column. However, as shown in Table 3, this peptide eluted much later than MEP, suggesting, that the bend in the a-helical structure of MEP imposed by Pro somehow perturbs the hydrophobic interactions of the MEP with the C-18 stationary phase, making it more "movable" within the column.…”

Amphipathic, α-helical antimicrobial peptides

“…It has in fact been shown that for model peptides, retention times are correlated to both structuring and amphipathicity, and directly correlated to the antimicrobial activity. 184 All the above considerations lead to the conclusion that the ability to structure into a well-defined, amphipathic ␣-helix is strongly correlated to antimicrobial activity. On the other hand, formation of a helical structure before membrane interaction may not be beneficial.…”

Application of modern reversed-phase peptide retention prediction algorithms to the Houghten and DeGraw dataset: Peptide helicity and its effect on prediction accuracy

“…This is not surprising as Pro is the well-known "helix-breaker" [20][21][22]. In this sequence framework, the prediction error is particularly high when substitution at position (6) is hydrophilic, i.e.…”