ChemInform Abstract: Rapid and Efficient Method for the Preparation of Fmoc‐Amino Acids Starting from 9‐Fluorenylmethanol.

KORTENAAR, P. B. W. TEN; DIJK, B. G. VAN; PEETERS, M.; RAABEN, B. J.; Adams, P. J.; Tesser, G. I.

doi:10.1002/chin.198639328

Cited by 3 publications

(3 citation statements)

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“…Deuterium-enriched alanines (Ala-d 4 or Fmoc-Ala-d 3 ) were purchased from Cambridge Isotope Laboratories (Andover, MA). The Ala-d 4 was manually derivatized with an Fmoc group as reported earlier 20 . Typically, two 2 H-labeled alanines were incorporated per peptide at different isotope abundance levels.…”

Section: Methodsmentioning

confidence: 99%

Accommodation of a Central Arginine in a Transmembrane Peptide by Changing the Placement of Anchor Residues

et al. 2012

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Both Trp and Arg in transmembrane protein domains make important interactions with lipids at the membrane/water interface, but at different depths. Derivatives of the designed peptide GWALP23, acetyl-GGALW5LALALALALALALW19LAGA-amide, with single Trp anchors, have proven useful for characterizing such interactions. Indeed previous work revealed quite different effects emanating from Arg substitutions at positions 12 and 14 within GWALP23, with the R12 peptide exhibiting multiple positions and orientations with respect to DOPC bilayer membranes (Vostrikov et al. [2010] J. Am. Chem. Soc. 132, 5803–5811). To gain further understanding of the multi-state behavior, we moved the Trp “anchor” residues to more outer positions 3 and 21 in GWALP23 itself, and in the R12 and R14 derivatives. The locations and orientations of the peptides with respect to lipid bilayer membranes of differing thickness were investigated by means of solid-state 2H NMR spectroscopy, using labeled alanines, and coarse-grained molecular dynamics simulations. Interestingly, relatively intense and narrow 2H resonances from selected backbone Cα deuterons were observed over quite narrow ranges of frequency and sample orientation. The backbone resonances reflect dynamic complexities and at the same time provide important contributions for the analysis of peptide transmembrane orientation. With the Trp3,21 anchors relatively far from the peptide and bilayer center, the results indicate significantly large apparent tilt angles, for example close to 30° for the new R12 and R14 peptides with respect to the bilayer normal of DLPC membranes. The R12 side chain indeed is “rescued” to a stable position, where it is accommodated within the transmembrane helix, when the Trp anchors are moved outward and to another face of the helix. At the same time, the R14 side chain of transmembrane GW3,21ALP23 also retains a stable favored position.

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Section: Methodsmentioning

confidence: 99%

Accommodation of a Central Arginine in a Transmembrane Peptide by Changing the Placement of Anchor Residues

et al. 2012

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“…Deuterated L-alanine-d 4 was obtained from Sigma Aldrich, and 9-fluorenylmethyloxycarbonyl (Fmoc) was used to protect its amino functionality as described by Ten Kortenaar et al (20) before being used in the synthesis. The peptides were isotopically labeled with one deuterium-labeled alanine residue at different positions in the transmembrane domain.…”

Section: Methodsmentioning

confidence: 99%

Influence of Flanking Residues on Tilt and Rotation Angles of Transmembrane Peptides in Lipid Bilayers. A Solid-State ²H NMR Study

et al. 2004

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To gain insight into the parameters that determine the arrangement of proteins in membranes, 2 H NMR experiments were performed to analyze tilt and rotation angles of membrane-spanning R-helical model peptides upon incorporation in diacylphosphatidylcholine bilayers with varying thickness. The peptides consisted of the sequence acetyl-GW 2 (LA) 8 LW 2 A-NH 2 (WALP23) and analogues thereof, in which the interfacial Trp residues were replaced by Lys (KALP23) and/or the hydrophobic sequence was replaced by Leu (WLP23 and KLP23). The peptides were synthesized with a single deuterium-labeled alanine at four different positions along the hydrophobic segment. For all peptides a small but systematic increase in tilt angle was observed upon decreasing the bilayer thickness. However, significantly larger tilt angles were obtained for the Lys-flanked KALP23 than for the Trp-flanked WALP23, suggesting that interfacial anchoring interactions of Trp may inhibit tilting. Increasing the hydrophobicity resulted in an increase in tilt angle for the Trp-flanked analogue only. For all peptides the maximum tilt angle obtained was remarkably small (less than 12°), suggesting that further tilting is inhibited, most likely due to unfavorable packing of lipids around a tilted helix. The results furthermore showed that the direction of tilt is determined almost exclusively by the flanking residues: Trp-and Lys-flanked peptides were found to have very different rotation angles, which were influenced significantly neither by hydrophobicity of the peptides nor by the extent of hydrophobic mismatch. Finally, very small changes in the side chain angles of the deuterated alanine probes were observed in Trp-flanked peptides, suggesting that these peptides may decrease their hydrophobic length to help them to adapt to thin membranes.Membrane proteins carry out many essential functions in cells. In the specific case of integral proteins, the membrane spanning parts of these molecules are in direct contact with acyl chains and headgroups of the surrounding lipids. Consequently, the lipid environment can modulate structure and dynamics and, hence, activity of membrane proteins (1). For example, changes in hydrophobic thickness of a lipid bilayer can affect the structure of an integral membrane protein by causing a hydrophobic mismatch between the bilayer thickness and the length of the membrane spanning parts of that protein (reviewed in refs 2-4). Understanding the molecular basis of how lipids influence membrane proteins requires precise information on structural properties of transmembrane segments of proteins and knowledge of how these properties are sensitive to protein and lipid composition. It is most suitable to perform such experiments on model membranes of synthetic lipids and transmembrane peptides, since the composition of both the lipids and the peptides can be systematically varied. Model membranes, made of phosphatidylcholine derivatives (PC) 1 and model transmembrane peptides, have already been extensively studied to inves...

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“…Unlabeled WALP23, KALP23, WLP23, and KLP23 peptides (for amino acid sequences, see Table 1) were synthesized using Fmoc/tBu solid-phase synthesis as described elsewhere for related KALP peptides (18). Deuteriumlabeled L-tryptophan (indole-d 5 , 98%) was purchased from Cambridge Isotopes Laboratories Inc. (Andover, MA), and Fmoc (9-fluorenylmethyloxycarbonyl) was used to protect its amino functionality as described by Ten Kortenaar et al (19) before being used in the peptide synthesis. The deuterium-labeled WALP23 peptides were synthesized according to the procedure described in ref 20 to prevent exchange of deuterons located at the indole ring against hydrogens in the presence of trifluoroacetic acid (TFA) (21).…”

Section: Methodsmentioning

confidence: 99%

Is There a Preferential Interaction between Cholesterol and Tryptophan Residues in Membrane Proteins?

et al. 2008

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Recently, several indications have been found that suggest a preferential interaction between cholesterol and tryptophan residues located near the membrane-water interface. The aim of this study was to investigate by direct methods how tryptophan and cholesterol interact with each other and what the possible consequences are for membrane organization. For this purpose, we used cholesterol-containing model membranes of dimyristoylphosphatidylcholine (DMPC) in which a transmembrane model peptide with flanking tryptophans [acetyl-GWW(LA) 8 LWWA-amide], called WALP23, was incorporated to mimic interfacial tryptophans of membrane proteins. These model systems were studied with two complementary methods.(1) Steady-state and time-resolved Förster resonance energy transfer (FRET) experiments employing the fluorescent cholesterol analogue dehydroergosterol (DHE) in combination with a competition experiment with cholesterol were used to obtain information about the distribution of cholesterol in the bilayer in the presence of WALP23. The results were consistent with a random distribution of cholesterol which indicates that cholesterol and interfacial tryptophans are not preferentially located next to each other in these bilayer systems. (2) Solid-state 2 H NMR experiments employing either deuterated cholesterol or indole ring-deuterated WALP23 peptides were performed to study the orientation and dynamics of both molecules. The results showed that the quadrupolar splittings of labeled cholesterol were not affected by an interaction with tryptophan-flanked peptides and, vice versa, that the quadrupolar splittings of labeled indole rings in WALP23 are not significantly influenced by addition of cholesterol to the bilayer. Therefore, both NMR and fluorescence spectroscopy results independently show that, at least in the model systems studied here, there is no evidence for a preferential interaction between cholesterol and tryptophans located at the bilayer interface.

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ChemInform Abstract: Rapid and Efficient Method for the Preparation of Fmoc‐Amino Acids Starting from 9‐Fluorenylmethanol.

Cited by 3 publications

References 1 publication

Accommodation of a Central Arginine in a Transmembrane Peptide by Changing the Placement of Anchor Residues

Accommodation of a Central Arginine in a Transmembrane Peptide by Changing the Placement of Anchor Residues

Influence of Flanking Residues on Tilt and Rotation Angles of Transmembrane Peptides in Lipid Bilayers. A Solid-State ²H NMR Study

Is There a Preferential Interaction between Cholesterol and Tryptophan Residues in Membrane Proteins?

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ChemInform Abstract: Rapid and Efficient Method for the Preparation of Fmoc‐Amino Acids Starting from 9‐Fluorenylmethanol.

Cited by 3 publications

References 1 publication

Accommodation of a Central Arginine in a Transmembrane Peptide by Changing the Placement of Anchor Residues

Accommodation of a Central Arginine in a Transmembrane Peptide by Changing the Placement of Anchor Residues

Influence of Flanking Residues on Tilt and Rotation Angles of Transmembrane Peptides in Lipid Bilayers. A Solid-State 2H NMR Study

Is There a Preferential Interaction between Cholesterol and Tryptophan Residues in Membrane Proteins?

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Influence of Flanking Residues on Tilt and Rotation Angles of Transmembrane Peptides in Lipid Bilayers. A Solid-State ²H NMR Study