Alain Blond

Structural changes of region 1-16 of the Alzheimer disease amyloid beta-peptide upon zinc binding and in vitro aging.

Amyloid deposits within the cerebral tissue constitute a characteristic lesion associated with Alzheimer disease. They mainly consist of the amyloid peptide Aβ and display an abnormal content in Zn2+ ions, together with many truncated, isomerized, and racemized forms of Aβ. The region 1-16 of Aβ can be considered the minimal zinc-binding domain and contains two aspartates subject to protein aging. The influence of zinc binding and protein aging related modifications on the conformation of this region of Aβ is of importance given the potentiality of this domain to constitute a therapeutic target, especially for immunization approaches. In this study, we determined from NMR data the solution structure of the Aβ-(1-16)-Zn2+ complex in aqueous solution at pH 6.5. The residues His6, His13, and His14 and the Glu11 carboxylate were identified as ligands that tetrahedrally coordinate the Zn(II) cation. In vitro aging experiments on Aβ-(1-16) led to the formation of truncated and isomerized species. The major isomer generated, Aβ-(1-16)-l-iso-Asp7, displayed a local conformational change in the His6-Ser8 region but kept a zinc binding propensity via a coordination mode involving l-iso-Asp7. These results are discussed here with regard to Aβ fibrillogenesis and the potentiality of the region 1-16 of Aβ to be used as a therapeutic target.

Siderophore peptide, a new type of post-translationally modified antibacterial peptide with potent activity

Microcin E492 (MccE492, 7886 Da), the 84-amino acid antimicrobial peptide from Klebsiella pneumoniae, was purified in a post-translationally modified form, MccE492m (8717 Da), from culture supernatants of either the recombinant Escherichia coli VCS257 strain harboring the pJAM229 plasmid or the K. pneumoniae RYC492 strain. Chymotrypsin digestion of MccE492m led to the MccE492m-(74–84) C-terminal fragment that carries the modification and that was analyzed by mass spectrometry and nuclear magnetic resonance at natural abundance. The 831-Da post-translational modification consists of a trimer of N-(2,3-dihydroxybenzoyl)-l-serine linked via a C-glycosidic linkage to a β-d-glucose moiety, itself linked to the MccE492m Ser-84-carboxyl through an O-glycosidic bond. This modification, which mimics a catechol-type siderophore, was shown to bind ferric ions by analysis of the collision-induced dissociation pattern obtained for MccE492m-(74–84) by electrospray ion trap mass spectrometry experiments in the presence of FeCl3. By using a series of wild-type and mutant isogenic strains, the three catechol-type siderophore receptors Fiu, Cir, and FepA were shown to be responsible for the recognition of MccE492m at the outer membrane of sensitive bacteria. Because MccE492m shows a broader spectrum of antibacterial activity and is more potent than MccE492, we propose that by increasing the microcin/receptor affinity, the modification leads to a better recognition and subsequently to a higher antimicrobial activity of the microcin. Therefore, MccE492m is the first member of a new class of antimicrobial peptides carrying a siderophore-like post-translational modification and showing potent activity, which we term siderophore-peptides.

Curcacycline B, a cyclic nonapeptide from Jatropha curcas enhancing rotamase activity of cyclophilin

The structure of curcacycline B (1), a cyclic nonapeptide isolated from Jatropha curcas latex was elucidated by combination of chemical degradation, LSIMS data and 2D NMR experiments. Curcacycline B was shown to enhance the rotamase activity of human cyclophilin B.

Pohlianins A, B and C, cyclic peptides from the laxes of Jatropha pohliana ssp. molissima

Abstract From the EtOAc extract of the latex of Jatropha pohliana (Euphorbiaceae), two cyclic heptapeptides, pohlianins A (1) and B (2) and one cyclic octapeptide, pohlianin C (3) were isolated by a multi-step chromatography procedure, including HPLC. Their structure were elucidated by chemical degradation, mass spectrometry, homonuclear and heteronuclear NMR experiments. Conformational analysis of these peptides was made by using NMR experiments and distance geometry calculations. Their antimalarial activities were examined.

Sequence Determinants Governing the Topology and Biological Activity of a Lasso Peptide, Microcin J25

Microcin J25 is a potent antibacterial peptide produced by Escherichia coli AY25. It displays a lasso structure, which consists of a knot involving an N‐terminal macrolactam ring through which the C‐terminal tail is threaded and sterically trapped. In this study, we rationally designed and performed site‐specific mutations in order to pinpoint the sequence determinants of the lasso topology. Structures of the resulting variants were analysed by a combination of methods (mass spectrometry, NMR spectroscopy, enzymatic digestion), and correlated to the antibacterial activity. The selected mutations resulted in the production of branched‐cyclic or lasso variants. The C‐terminal residues below the ring (Tyr20, Gly21) and the size of the macrolactam ring were revealed to be critical for both the lasso scaffold and bioactivity, while shortening the loop region (Tyr9–Ser18) or extending the C‐terminal tail below the ring did not alter the lasso structure, but differentially affected the antibacterial activity. These results provide new insights for the bioengineering of antibacterial agents using a lasso peptide as template.

Mahafacyclin A, a cyclic heptapeptide from Jatropha mahafalensis exhibiting β-bulge conformation

Abstract The structure of mahafacyclin A ( 1 ), a cyclic heptapeptide isolated from Jatropha mahafalensis latex was elucidated by a combination of chemical degradation, LSIMS data and 2D NMR experiments. Mahafacyclin A solution conformation was shown to have β-bulge characteristics.

Characterization of Sviceucin from Streptomyces Provides Insight into Enzyme Exchangeability and Disulfide Bond Formation in Lasso Peptides.

Lasso peptides are bacterial ribosomally synthesized and post-translationally modified peptides. They have sparked increasing interest in peptide-based drug development because of their compact, interlocked structure, which offers superior stability and protein-binding capacity. Disulfide bond-containing lasso peptides are rare and exhibit highly sought-after activities. In an effort to expand the repertoire of such molecules, we heterologously expressed, in Streptomyces coelicolor, the gene cluster encoding sviceucin, a type I lasso peptide with two disulfide bridges originating from Streptomyces sviceus, which allowed it to be fully characterized. Sviceucin and its reduced forms were characterized by mass spectrometry and peptidase digestion. The three-dimensional structure of sviceucin was determined using NMR. Sviceucin displayed antimicrobial activity selectively against Gram-positive bacteria and inhibition of fsr quorum sensing in Enterococcus faecalis. This study adds sviceucin to the type I lasso peptide family as a new representative. Moreover, new clusters encoding disulfide-bond containing lasso peptides from Actinobacteria were identified by genome mining. Genetic and functional analyses revealed that the formation of disulfide bonds in sviceucin does not require a pathway-encoded thiol-disulfide oxidoreductase. Most importantly, we demonstrated the functional exchangeability of the sviceucin and microcin J25 (a non-disulfide-bridged lasso peptide) macrolactam synthetases in vitro, highlighting the potential of hybrid lasso synthetases in lasso peptide engineering.

LUTOSIDE : AN ACYL-1-(ACYL-6'-MANNOBIOSYL)-3-GLYCEROL ISOLATED FROM THE SPONGE-ASSOCIATED BACTERIUM MICROCOCCUS LUTEUS

Abstract Lutoside, an unusual acyl-1-(acyl-6′-mannobiosyl)-3-glycerol 1 was isolated from the sponge-associated bacterial strain Microccocus luteus. Sructure elucidation was performed by sprectroscopic analysis and chemical transformations.

16β-Hydroxy-5α-cholestane-3,6-dione, a novel cytotoxic oxysterol from the red alga Jania rubens

Abstract A new cytotoxic oxysterol, 16β-hydroxy-5α-cholestane-3,6-dione was isolated from the red alga Jania rubens. Its structure was established by spectroscopic method. The ID50 value was 0.5 μg/mL.

Isolation, structure and synthesis of mahafacyclin B, a cyclic heptapeptide from the latex of Jatropha mahafalensis

Mahafacyclin B is a cyclic heptapeptide with antimalarial activity isolated from the latex of Jatropha mahafalensis. The structure is elucidated by chemical degradation, tandem mass spectrometry, homo- and heteronuclear NMR experiments, and confirmed by synthesis.