Retention behaviour of a template-assembled synthetic protein and its amphiphilic building blocks on reversed-phase columns

Steiner,; Schär, Martin; Ko, Börnsen; Mutter, Manfred

doi:10.1016/0021-9673(91)80023-a

Cited by 42 publications

(18 citation statements)

References 12 publications

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“…In an α‐helical conformation, this designer peptide is amphiphilic along the length of the helix 108. In the original contribution it was stated that cellular uptake of this CPP occurs primarly through nonendocytic uptake and depends primarily on helical amphipathicity 109.…”

Section: Biophysics and Cell Biology Case By Casementioning

confidence: 99%

Break on through to the Other Side—Biophysics and Cell Biology Shed Light on Cell‐Penetrating Peptides

et al. 2005

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Cell-penetrating peptides (CPPs) have become widely used vectors for the cellular import of molecules in basic and applied biomedical research. Despite the broad acceptance of these molecules as molecular carriers, the details of the mode of cellular internalization and membrane permeation remain elusive. Within the last two years endocytosis has been demonstrated to be a route of uptake shared by several CPPs. These findings had a significant impact on CPP research. State-of-the-art cell biology is now required to advance the understanding of the intracellular fate of the CPP and cargo molecules. Owing to their presumed ability to cross lipid bilayers, CPPs also represent highly interesting objects of biophysical research. Numerous studies have investigated structure-activity relationships of CPPs with respect to their ability to bind to a lipid bilayer or to cross this barrier. Endocytosis route only relocates the membrane permeation from the cell surface to endocytic compartments. Therefore, biophysical experiments are key to a mechanistic molecular understanding of the cellular uptake of CPPs. However, biophysical investigations have to consider the molecular environment encountered by a peptide inside and outside a cell. In this contribution we will review biophysical and cell-biology data obtained for several prominent CPPs. Furthermore, we will summarize recent findings on the cell-penetrating characteristics of antimicrobial peptides and the antimicrobial properties of CPPs. Peptides of both groups have overlapping characteristics. Therefore, both fields may greatly benefit from each other. The review will conclude with a perspective of how biophysics and cell biology may synergize even more efficiently in the future.

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Section: Biophysics and Cell Biology Case By Casementioning

confidence: 99%

Break on through to the Other Side—Biophysics and Cell Biology Shed Light on Cell‐Penetrating Peptides

et al. 2005

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“…Thus, if a molecule becomes helical on binding and contains a preferred binding domain (i.e., the non-polar face of an amphipathic α-helix), the binding of this domain to the hydrophobic stationary phase will be greater than if it were binding as a random coil or as a non-amphipathic α-helix [24,28], showing significant deviation from expected retention behaviour based on amino acid composition alone. Significant differences in retention time between α-helical peptides of the same composition but different sequence (SCDS) were also demonstrated by Houghten's group [29,30].…”

Section: (Iii) Sequence Dependent Effectsmentioning

confidence: 99%

“…All peptides have a minimum of a single positive charge to ensure peptide solubility at pH 2. The four hydrophobes in the peptide sequences 1, 2, 3 and 4 (2 Ala, 1 Val and 1 Leu) and peptide 5 (4 Leu) were distributed throughout the sequence to ensure no clustering of hydrophobes and subsequent creation of a preferred hydrophobic binding domain [24,28]. The sequences of Peptide 1 and Peptide 2 are identical (Table 1), differing only in the end-group (C α -carboxyl or C α -amide group).…”

Section: Design Of Model Peptidesmentioning

confidence: 99%

Requirements for prediction of peptide retention time in reversed-phase high-performance liquid chromatography: Hydrophilicity/hydrophobicity of side-chains at the N- and C-termini of peptides are dramatically affected by the end-groups and location

Tripet

Cepeniene

Kovacs

et al. 2007

Journal of Chromatography A

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The value of reversed-phase high-performance liquid chromatography (RP-HPLC) and the field of proteomics would be greatly enhanced by accurate prediction of retention times of peptides of known composition. The present study investigates the hydrophilicity/hydrophobicity of amino acid sidechains at the N-and C-termini of peptides while varying the functional end-groups at the termini. We substituted all 20 naturally occurring amino acids at the N-and C-termini of a model peptide sequence, where the functional end-groups were N α -acetyl-X-and N α -amino-X-at the N-terminus and -X-C α -carboxyl and -X-C α -amide at the C-terminus. Amino acid coefficients were subsequently derived from the RP-HPLC retention behaviour of these peptides and compared to each other as well as to coefficients determined in the centre of the peptide chain (internal coefficients). Coefficients generated from residues substituted at the C-terminus differed most (> 2.5 min between the -X-C α -carboxyl and -X-C α -amide peptide series) for hydrophobic side-chains. A similar result was seen for the N α -acetyl-X-and N α -amino-X-peptide series, where the largest differences in coefficient values (> 2 min) were observed for hydrophobic peptides. Coefficients derived from substitutions at the Cterminus for hydrophobic amino acids were dramatically different compared to internal coefficients for hydrophobic side-chains, ranging from 17.1 min for Trp to 4.8 min for Cys. In contrast, coefficients derived from substitutions at the N-terminus showed relatively small differences from the internal coefficients. Subsequent prediction of peptide retention time, within an error of just 0.4 min, was achieved by a predictive algorithm using a combination of internal coefficients and a weighted coefficient for the C-terminal residue.

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“…Broadening of HPLC peaks was pronounced for large carboprotein structures when analyzing them on a C18 RP-HPLC column; substituting a C18 for a C4 RP column significantly sharpened the peaks. We attribute this phenomenon to the presence of equilibria between folded and unfolded bundle structures and their different affinity for the column material [63,64]. It seems unlikely that it is due to the presence of E/Z isomers.…”

Section: The Second Generationmentioning

confidence: 89%

Carbohydrates as templates for control of distance-geometry in de novo-designed proteins

Jensen¹,

Brask

2002

CMLS, Cell. Mol. Life Sci.

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An understanding of very complex natural systems can often only be achieved through detailed studies of systems with a reduced complexity. Thus, de novo design of proteins allows the study of fundamental forces determining protein folding and stability, as well as protein-protein interactions, by analyses of protein models of structural motifs. In addition, de novo design may lead to new biomimetic molecules with novel properties. In a synthetic approach to achieve structural economy, rigid templates, sometimes called topological scaffolds, have been used to connect secondary-structure elements, most notably alpha-helices. By positioning the helices on the template, the unfavorable entropy of protein folding is reduced. In a novel class of chimeric molecules called carboproteins, carbohydrates are used as templates for de novo design of protein models. Recently, a strategy relying on chemoselective ligation of C-terminal peptide aldehydes to tetra-aminooxy functionalized monosaccharides has provided 7-kDa 4-alpha-helix bundle carboproteins.

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Retention behaviour of a template-assembled synthetic protein and its amphiphilic building blocks on reversed-phase columns

Cited by 42 publications

References 12 publications

Break on through to the Other Side—Biophysics and Cell Biology Shed Light on Cell‐Penetrating Peptides

Break on through to the Other Side—Biophysics and Cell Biology Shed Light on Cell‐Penetrating Peptides

Requirements for prediction of peptide retention time in reversed-phase high-performance liquid chromatography: Hydrophilicity/hydrophobicity of side-chains at the N- and C-termini of peptides are dramatically affected by the end-groups and location

Carbohydrates as templates for control of distance-geometry in de novo-designed proteins

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