Quirinus B. Broxterman

The First Water-Soluble 310-Helical Peptides

Two water-soluble 3(10)-helical peptides are synthesized and fully characterized for the first time. The sequence of these terminally blocked heptamers comprises two residues of the Calpha-trisubstituted alpha-amino acid 2-amino-3-[1-(1,4,7-triazacyclononyl)]propanoic acid and five residues of a Calpha-tetrasubstituted alpha-amino acid (either alpha-aminoisobutyric acid or isovaline). Using CD and NMR techniques we were able to show that both heptapeptides are well structured in water, and that the type of conformation adopted is indeed the ternary 3(10)-helix.

Enzymatic resolution of α,α-disubstituted α-amino acid esters and amides

The scope and limitations of the enzymatic resolution of α,α-disubstituted α-amino acid amides by an amino acid amidase from Mycobacterium neoaurum and of the corresponding ethyl esteis with Pig liver estetase (PLE) have been studied. Moderate enantiomeric excesses were obtained with PLE, with only a narrow substrate specificity. Mycobacterium neoaurum on the contrary yields a broad range of S-α,α-disubstituted α-amino acids 1 and the corresponding R-amides 2.

A peptide template as an allosteric supramolecular catalyst for the cleavage of phosphate esters

The heptapeptide H-Iva-Api-Iva-ATANP-Iva-Api-Iva-NHCH3 (P1a), where Iva is (S)-isovaline, Api is 4-amino-4-carboxypiperidine, and ATANP is (S)-2-amino-3-[1-(1,4,7-triazacyclononane)]propanoic acid, has been synthesized. Its conformation in aqueous solution is essentially that of a 310-helix. By connecting three copies of P1a to a functionalized Tris(2-aminoethyl)amine (Tren) platform a new peptide template, [T(P1)3], was obtained. This molecule is able to bind up to four metal ions (CuII or ZnII): one in the Tren subsite and three in the azacyclononane subunits. The binding of the metals to the Tren platform induces a change from an open to a closed conformation in which the three short, helical peptides are aligned in a parallel manner with the azacyclonane units pointing inward within the pseudocavity they define. T(P1)3 shows a peculiar behavior in the transphosphorylation of phosphate esters; the tetrazinc complex is a catalyst of the cleavage of 2-hydroxypropyl-p-nitrophenyl phosphate (HPNP), whereas the free ligand is a catalyst of the cleavage of an oligomeric RNA sequence with selectivity for pyrimidine bases. In the case of HPNP, ZnII acts as a positive allosteric effector by enhancing the catalytic efficiency of the system. In the case of the polyanionic RNA substrate, ZnII switches off the activity, thus behaving as a negative allosteric regulator. It is suggested that the opposite behavior of the catalyst induced by ZnII is associated with the change of conformation of the Tren platform, and consequently of the relative spatial disposition of the three linked peptides, that occurs after binding of the metal ion.

Nitroxyl peptides as catalysts of enantioselective oxidations.

The achiral, nitroxyl-containing alpha-amino acid TOAC (TOAC = 2,2,6,6-tetramethylpiperidine-1-oxyl-4-amino-4-carboxylic acid), in combination with the chiral alpha-amino acid C(alpha)-methyl valine [(alphaMe)Val], was used to prepare short peptides (from di- to hexa-) that induced the enantioselective oxidation of racemic 1-phenylethanol to acetophenone. The best catalyst was an N(alpha)-acylated dipeptide alkylamide with the -TOAC-(alphaMe)Val- sequence folded in a stable, intramolecularly hydrogen-bonded beta-turn conformation with large, lipophilic (hydrophobic) N- and C-terminal blocking groups. We rationalized our findings by proposing models for the diastereomeric intermediates between (R)-[and (S)]-1-phenylethanol and the catalyst Fmoc-TOAC-L-(alphaMe)Val-NHiPr, based on the X-ray diffraction structure of the latter.

High-performance liquid chromatographic separation of the enantiomers of unusual α-amino acid analogues

The direct and indirect stereochemical resolution of the enantiomers of ring- and alpha-methyl-substituted phenylalanines and phenylalanine amides was attempted by high-performance liquid chromatographic methods. The direct separation was carried out on two chiral stationary phases, the crown-ether-based Crownpak CR(+), and the teicoplanin-based Chirobiotic T, while the indirect resolution was performed by applying pre-column derivatization with 2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl isothiocyanate (GITC) and Nalpha-(2,4-dinitro-5-fluorophenyl)-L-alanine amide (Marfeys reagent, FDAA). The Chirobiotic T column was efficient in the separation of ring- and alpha-methyl-substituted phenylalanine analogues, but was ineffective for the amides of these analogues. The Crownpak CR(+) column separated the ring-substituted phenylalanines and amides, whereas the alpha-methylated analogues were coeluted. Of the two indirect methods, GITC derivatization seemed more effective than FDAA derivatization.

Aspartame dipeptide analogues: effect of number of side-chain methylene group spacers and Cα-methylation in the second position

Abstract Our model of the active site of the sweet taste receptor is shown to be consistent with the aspartame analogues in which the L-Phe 2 residue is replaced by L-(αMe)Phg, L-(αMe)Phe or L-(αMe)Hph. The analogues containing either the first or the third C α -methylated, phenyl-containing residue in the second position of the dipeptide were synthesized and found to be approximately as sweet as aspartame itselfand its L-(αMe)Phe 2 analogue.

First homo‐peptides undergoing a reversible 310‐helix/α‐helix transition: Critical main‐chain length

The difference in length between the more elongated peptide 310‐helix and the more compact α‐helix is about 0.4 Å/residue. This property makes the 310‐/α‐helix reversible conversion very promising as a molecular switching tool between the N‐ and C‐terminal functions of a peptide backbone. In this work, using homo‐peptides of various main‐chain length, all based on the strongly helicogenic, Cα‐tetrasubstituted α‐amino acid Cα‐methyl‐L‐valine, we show that a well defined, solvent controlled, reversible 310‐/α‐helix transition takes place even in a homo‐oligomer as short as a terminally blocked hexapeptide. Homo‐peptide sequences blocked as a urethane or an acetamide at the N‐terminus and as a methyl ester or an N‐alkyl amide at the C‐terminus are all appropriate. The nature of the occurring helical species in the various solvents tested was assessed by electronic or vibrational circular dichroism.

New aspartame-like sweeteners containing L-(αMe)Phe

Abstract The [L-(αMe) Phe] 2 -analogue of aspartame was synthesized and analyzed by X-ray diffraction. This compound is as sweet as aspartame itself but far more stable at pH=4. Several N-protected analogues were synthesized. The N-formylcarbamoyl [L-(αMe Phe)] 2 -aspartame analogue is also sweet. The compounds fit well within the sweet perception model as developed by Temussi, Toniolo and coworkers.

Chiral, fully extended helical peptides.

The synthesis of the N-protected (blocked) homo-peptide esters from the chiral Cα-ethyl, Cα-n-pentylglycine was performed in solution to the hexapeptide level. The conformational propensity exhibited by these oligomers in chloroform solution and in the crystal state was assessed by use of FTIR absorption, NMR, and X-ray diffraction. The results indicated that fully extended helical structures (2.05-helices) are overwhelmingly adopted irrespective of the peptide main-chain length. This oligomeric series is of great interest as it is characterized by the longest Ciα,…, Ci+1α (per residue) separation achievable in the class of chiral, rigid, helical peptide spacers based on α-amino acids.

Microbiological transformations 50: selection of epoxide hydrolases for enzymatic resolution of 2-, 3- or 4-pyridyloxirane

Abstract A study aimed to select efficient epoxide hydrolases (EHs) allowing to achieve the enzymatic resolution of 2-, 3- and 4-pyridyloxirane (1–3) has been achieved, using 2-pyridyloxirane 1 as test substrate. Five thus selected EH-sources that showed interesting enantioselectivity were looked at in more detail for the conversion of 1–3.