Miroslaw T. Leplawy

C-terminally shortened alamethicin on templates: influence of the linkers on conductances

In order to test the influence of chemical modifications designed to allow covalent coupling of channel-forming peptide motifs into variable sized oligomers, a series of alamethicin derivatives was prepared. The building block encompassing the N-terminal 1-17 residues of alamethicin behaved normally in the conductance assay on planar lipid bilayers, albeit at higher concentration and with a slightly reduced voltage-dependence. A linker Ac-K-OCH(2)C(6)H(4)CH(3)p attached via the epsilon amino group of lysine to the C-terminus of alamethicin(1-17) increased membrane affinity. The latter was further enhanced in a dimer and a tetramer in which alamethicin(1-17) chains were tethered to di- or tetra-lysine linkers, respectively, but macroscopic current-voltage curves displayed much reduced voltage-dependencies and reversed hysteresis. An usual behaviour with high voltage-dependence was restored with the modified dimer of alamethicin(1-17) in which alanine separated the two consecutive lysine residues in the linker. Of special interest was the development of a negative resistance branch in macroscopic current-voltage curves for low concentrations of this dimer with the more flexible linker. Single channel events displayed only one single open state with fast kinetics and whose conductance matches that of the alamethicin heptamer or octamer.

Preferred structures of constrained peptides from achiral α,α-dialkyiated glycyl residues with acyclic side chains

Conformational energy computations of the monopeptides from three achiral α,α-dialkylated glycyl residues with acyclic side chains (namely α,α-dimethyl-; α,α-diethyl-; and α, α-di-n-propylglycines) are reported as a function of the relevant N-Cα-C′ bond angle. In parallel, experimental studies were performed in the solid state (infrared absorption and X-ray diffraction) and in solution (infrared absorption and proton magnetic resonance) on the corresponding protected homo-peptide series (the former series to the dodecamer, the other two series to the pentamers). The results obtained unequivocally indicate that the preference from a helical to a fully extended conformation increases as side-chain bulkiness increases. The longest homo-peptides from α,α-dimethylglycine form stable 310-helices. A picture of the mode of self-association of the helical structures has also been determined. The results of the theoretical analyses fit well with the experimentally observed conformational properties in the solid state and in chloroform solution.

Effect of α-hydroxymethylserine residue on binding ability of oligopeptides to Cu2+ ions☆

Abstract Potentiometric and spectroscopic data have shown that α-hydroxymethyl-L-serine residue may play an important role in the coordination of tripeptides to Cu 2+ ions. The alcoholic groups in the amino acid side chain are very effective in the stabilization of the complexes formed. It is also likely that at basic solutions, the alcoholic group may dissociate proton and bind to metal ion directly via the O − oxygen. The increase in stability is up to 3.7 orders of magnitude when compared to simple tri-Ala containing systems.

Copper(II) complexes of deltakephalin and its analogues containing α-hydroxymethylserine

Abstract Potentiometric and spectroscopic data have shown that deltakephalin and its derivatives containing the α-hydroxymethylserine (HmS) residue in the second and in the second and sixth positions form the same series of complexes: CuHL, CuL, CuH−1L, CuH−2L and CuH−3L (1N to 4N binding mode). The alcoholic groups in the amino acid side chain are very effective in the stabilization of the complexes formed (the protonation corrected stability constants, logxa0K*, values of 4N complexes are −22.55 and −21.15 for deltakephalin and its derivative containing the HmS residue in the second and sixth positions, respectively). There is no evidence for the direct involvement of alcoholic groups of HmS in binding of Cu2+ ions and there is no indication of binding through the tyrosine-phenolate oxygen atoms, either.

Introduction of α-hydroxymethylserine residues in a peptide sequence results in the strongest peptidic, albumin-like, copper(II) chelator known to date

A tripetide amide HmS-HmS-His-NH2 is the strongest peptidic CuII chelator known to date, due to the steric shielding of the chelate plane as well as electronic effects.

α-Hydroxymethylserine as a peptide building block: synthetic and structural aspects

A synthetic methodology has been developed for peptide bond formation with α‐hydroxmethylserine as the carboxyl or amino component and also for the preparation of homo‐sequences. The key intermediate, O,O‐protected α‐hydroxymethylserine in the form of an isopropylidene derivative, is easily accessible and represents the first example of a heterocyclic Cα,α‐disubstituted amino acid containing an 1,3‐dioxane ring. The use of this intermediate facilitates protection of the sterically hindered amino and carboxyl groups and is advantageous for the coupling and deprotection steps. X‐ray structure determination of Z‐HmS(Ipr)–Ala–OMe revealed that the two crystallographically independent molecules present in the asymmetric unit adopt an S‐shaped conformation. In the one molecule the achiral HmS(Ipr) residue has the torsion angle values (ϕ==61.4°,ψ=40.8°) in the left‐handed helical region of the Ramachandran map, while in the second molecule the negative torsion angles (ϕ=−60.1°, ψ=–44.4°) are associated with the right‐handed helix.

IMPACT OF α-HYDROXYMETHYLSERINE RESIDUES ON BINDING ABILITY OF DIPEPTIDES TOWARDS CUII IONS

Abstract Potentiometric and spectroscopic studies performed on the binding ability of dipeptides containing α-hydroxymethylserine have shown that presence of the hydroxylmethyl substituent on the α-carbon changes considerably stability constants and binding modes. The indirect and direct involvement of the additional hydroxyl group is likely and this protects metal-peptide complexes against hydrolysis in very basic solutions.

Copper(II) and oxovanadium(IV) complexes of α-hydroxymethylserine

SummaryPotentiometric and spectroscopic measurements were used to characterize the binding ability of α-hydroxy-methylserine (Hms) with copper(II) and oxovanadium(IV) ions. The ligand was found to be generally a more efficient chelating agent than serine. Both of the deprotonated hydroxyl groups of Hms can be involved in coordination to vanadyl ions, whereas copper(II) binds in the same way as with serine.

Peptides derived from α-hydroxymethylserine: Aspects of solid-phase synthesis

The solid-phase synthesis of peptides derived from the sterically hindered α-hydroxymethylserine (HmS) was investigated. The acid-sensitive,O,O-isopropylidene (Ipr) protection of HmS is compatible with the Fmoc chemistry, represented here by the Fmoc-HmS(Ipr)-OH and Fmoc-HmS(Ipr)-F derivatives. Three analogs of the opioid pentapeptide DADLE with a single or two consecutive HmS residue(s) were synthesized using Wang resin as the solid support. The HATU method has been shown to effectively accomplish ‘difficult’ couplings with the HmS(Ipr) residue. Wang resin is not suitable, for the synthesis of sequences with a C-terminal HmS because of the easy formation of the diketopiperazine resulting from the cyclization of the susceptible dipeptide sequence AA-HmS(Ipr) bound to the resin. A further drawback of the Wang resin methodology is the increased danger of the undersired N→O-acyl shift, when long-lasting acidic cleavage is applied. These side reactions are totally suppressed when the 2-chlorotrityl polystyrene is used as a solid support. The mild conditions (AcOH/TFE/DCM) applied for the peptide detachment from this resin do not affect the Ipr protection, affording highly pure fragments with HmS(Ipr) residues suitable for post-cleavage condensation, cyclization or controlled side-chain deprotection. This approach is documented by the efficient synthesis of linear and cyclic analogs of the opioid hexapeptide DTLET containing two residues of HmS or HmS(Ipr) in positions 2 and 6.

Copper(II) complexes of (R,S)-α-hydroxymethylornithine and its Nδ-benzoyl derivative

Abstract Complex formation between copper(II) and ( R,S )-α-hydroxymethylornithine of ( R,S )- N δ -benzoyl-α-hydroxymethylornithine was studied in aqueous solution by potentiometric and spectroscopic (electron paramagnetic resonance and electronic absorption) techniques. The results show that the α-hydroxymethyl derivatives of ornithine are coordinated through the alcoholic group to the copper(II) ion in basic solution. Deprotonation and coordination of the α-hydroxymethyl group occurs and yields species with the amino and/or car☐ylato groups also bound to the metal ion.