Henri Pepermans

1H NMR study of the interaction of N,N′,N″-triacetyl chitotriose with Ac-AMP2, a sugar binding antimicrobial protein isolated from Amaranthus caudatus

The interaction between Ac‐AMP2, a lectin‐like small protein with antimicrobial and antifungal activity isolated from Amaranthus caudatus, and N,N′,N″‐triacetyl chitotriose was studied using 1H NMR spectroscopy. Changes in chemical shift and line width upon increasing concentration of N,N′,N″‐triacetyl chitotriose to Ac‐AMP2 solutions at pH 6.9 and 2.4 were used to determine the interaction site and the association constant K a. The most pronounced shifts occur mainly in the C‐terminal half of the sequence. They involve the aromatic residues Phe18, Tyr20 and Tyr27 together with their surrounding residues, as well as the N‐terminal Val‐Gly‐Glu segment. Several NOEs between Ac‐AMP2 and the N,N′,N″‐triacetyl chitotriose resonances are reported.

Tryptophan mediated photoreduction of disulfide bond causes unusual fluorescence behaviour of Fusarium solani pisi cutinase

The fluorescence signal of the single tryptophan residue (Trp69) of Fusarium solani pisi cutinase is highly quenched. However, prolonged irradiation of the enzyme in the tryptophan absorption band causes an increase of the tryptophan fluorescence quantum yield by an order of magnitude. By using a combination of NMR spectroscopy and chemical detection of free thiol groups with a sulfhydryl reagent we could unambiguously show that the unusual fluorescence behaviour of Trp69 in cutinase is caused by the breaking of the disulfide bond between Cys31 and Cys109 upon irradiation, while the amide‐aromatic hydrogen bond between Ala32 and Trp69 remains intact. This is the first example of tryptophan mediated photoreduction of a disulfide bond in proteins.

Evidence for the bioactive conformation in a cyclic hexapeptide analogue of somatostatin containing a cis-peptide bond mimic

N-methyl- alpha -benzyl-o-aminomethylphenylacetic acid was incorporated into a cyclic somatostatin analogue in order to mimic a cis-peptide bond configuration. The high biological potency of one of the isomers of the cyclic peptide strongly argues in favour of the proposed cis-configuration of the peptide bond at that position in the parent peptide. This represents the first cis-peptide bond mimic which has high biological activity.

Synthesis and conformational analysis of a series of galactosyl enkephalin analogues showing high analgesic activity

Two galactosyl derivatives of [DMet2,Pro5] enkephalin‐amide (compound 1), namely [DMet2,Pro5] enkephalin [N1.5‐beta‐D‐galactopyranosyl] amide (compound 2) and O1.5‐(beta‐D‐galactopyranosyl) [DMet2,Hyp5] enkephalin‐amide (compound 3) have been synthesized. Such glycosylpeptides have been shown to be extremely potent analgesic agonists. The conformational analysis of these three compounds in DMSO‐d6 solution has been carried out using two‐dimensional NMR methods. Both the parent compound (1) and the beta N‐galactosyl derivative (2) show similar NMR parameters which are consistent with fairly rigid beta‐strands at both the N‐terminus and C‐terminus, connected by a glycine residue that displays a mixture between multiple conformational states. Thus, although the beta N‐galactosyl derivative (2) has been shown to be significantly more potent than the parent compound (1) in the tail immersion and paw pressure tests of analgesia, no correlation can be established between the conformation of (1) and (2) in DMSO and the difference in analgesic activity. In contrast, important conformational differences with respect to (1) and (2) have been detected in the beta O‐galactosyl derivative (3). In this case, only one of the likely conformations for (1) and (2) are consistent with the experimental data. These data show that the position of the galactose residue in compound (3) causes Gly3 to loose flexibility leading to a more rigid folded conformation. Such a change in conformation could be related to the difference in analgesic activity between (2) and (3).

Identification and sequencing of sugars in an acetylated saponin of Blighia welwitschii by n.m.r. spectroscopy

Abstract High-resolution 2D 1 H-n.m.r. spectroscopy has been used to establish the structure of an acetylated pentasaccharide saponin from the fruit of Blighia welwitschii as acetylated 3β- O -[β- d -Xyl p -(1→3)-α- l -Ara p -(1→4)-β- d -Glc p -(1→3)-α- l -Rha p -(1→2)-α- l -Ara p ]hederagenin .