Thesis (PhD (Chemistry and Polymer Science)--University of Stellenbosch, 2006. / Synthetic peptides derived from the active core of a natural antimicrobial peptide were used as a template for the design of novel bolaamphiphilic peptides and hybrid molecules. The amphiphilic character of the original compounds was modified by using non-natural amino acids (AAs) – such as ω-AA – and varying the hydrophobic content. The outcomes of these modifications were studied focusing on structural and biological properties. Because of the bolaamphiphilic character, the alternation of polar and non-polar AAs and the use of hydrophobic AAs such as tyrosine and leucine, these novel molecules were designed to undergo self-assembly in response to certain stimuli (e.g. a pH increase). This significant property was investigated by means of different tools, such as fluorescence measurements, electron microscopy (EM), Fourier transform infrared spectroscopy (FT-IR) and circular dichroism (CD). By using fluorescence it was possible to determine the critical aggregation concentration (CAC) of the new compounds. Differences in amino acid composition, which were reflected into diverse secondary structures and hydrophobicity (H), resulted in different CAC values and aggregation profiles. The data were consistent with the literature and showed that (i) the aggregation of these basic compounds was triggered by a pH increase, (ii) the use of hydrophobic AA highly augmented the self-assembly tendency while (iii) the presence of proline strongly reduced it. EM revealed the morphology of the peptide assemblies: microtubes and microvesicles were identified and characterised by dimensions of 500 nm to 2 μm. The presence of 3-way junctions and vesicles budding out of the microtubes demonstrated that the self-assembly is a dynamic process. The aggregation was confirmed by FT-IR spectroscopy, by studying the dried peptide assemblies and the significant spectral signs the process left, especially in the amide II envelope. The relationship between hydrophobicity and self-assembly was expanded by experimentally and theoretically determining the hydrophobic content of the novel bolaamphiphiles. Data from liquid chromatography and computational calculations (two common ways used to determine the hydrophobicity of a given molecule) correlated well with the tendency to self-assemble, as expressed by CAC values. Importantly, some structural parameters (such as the presence of β-turn induced by proline) also showed significant influence on the aggregation, highly limiting the role of the peptides’ hydrophobicity. These novel peptide bolaamphiphiles displayed a very low haemolytic action and retained some antimicrobial activity at high concentrations against both Gram-positive and -negative bacteria. Unfortunately, the activity was greatly reduced at low concentrations, as clearly demonstrated by the use of two antimicrobial tests. The inability to provoke cell lysis was also evident when using liposomes mimicking a negative bacterial membrane. The loss of activity is possibly related to the modifications of the three-dimensional structure caused by the use of ω-AA and proline, which strongly alter the secondary structure. The results of this study were valuable in terms of understanding the relationships between self-assembly and structural parameters, such as AA compositions, hydrophobicity and secondary structure. Possible applications of the synthesised compounds were however limited as a result of the loss of the biological activity at low concentrations.

京都大学 / 新制・課程博士 / 博士(工学) / 甲第25011号 / 工博第5188号 / 新制||工||1990(附属図書館) / 京都大学大学院工学研究科分子工学専攻 / (主査)教授 SIVANIAH Easan, 教授 今堀 博, 教授 寺村 謙太郎 / 学位規則第4条第1項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DFAM

Ces travaux de thèse portent sur l’étude chimique d’espèces marines échantillonnées en Méditerranée. Dans le cadre du projet VALBIM, co-financé par la région PACA et en étroite collaboration avec la société BioPreserv à Grasse mais également avec la Mycoteca Universitatis Taurinensis (MUT) de l’Université de Turin, nous avons isolé et caractérisé 21 molécules dont 14 se sont avérées nouvelles. L’étude de l’algue Caulerpa taxifolia a conduit à la caulerpényne et à quatre nouveaux dérivés de l’acide pyruvique sulfatés. Les travaux sur l’ascidie Polysyncraton sp. ont conduit aux bolascidines A-D, quatre nouveaux métabolites de type bolaamphiphiles dont il n’existe pas à notre connaissance d’équivalent naturel ou synthétique. La première étude chimique de l’éponge Hexadella racovitzai a conduit à deux dérivés de la bromotyrosine, la psammapline A et une nouvelle molécule, la 4-Osulfatocyclobispsammapline A. Par ailleurs, trois nouvelles crambescines C1, conjointement avec une crambescine A2 déjà répertoriée, ont été isolées de Crambe crambe. L’étude du champignon marin Stachybotrys chartarum, isolé de l’éponge Aplysina cavernicola, a conduit à la satratoxine H et à deux nouveau dérivé, la 2,3-dihydrosatratoxine H et l’épi-chartarutine G, ainsi qu’à trois dérivés stachybotrylactame déjà répertoriés dans la bibliographie. Les différents métabolites ont été évalués pour leurs propriétés antibactériennes et cytotoxiques. Ces travaux ont bénéficié de nouvelles approches analytiques (calculs des spectres de dichroïsme circulaire électroniques par TDDFT, réseaux moléculaires) permettant de renforcer l’intérêt des études liées à l’écologie chimique et à la recherche de nouvelles molécules bioactives potentiellement valorisables dans différents domaines. / This PhD work focuses on the chemical study of marine species sampled in the Mediterranean Sea. As part of the VALBIM project, cofinanced by the region PACA and in close collaboration with the SME BioPreserv in Grasse but also with the Mycoteca Universitatis Taurinensis (MUT) of the University of Turin, we have isolated and characterized 21 molecules in total.14 of them were never reported before. The study of the seaweed Caulerpa taxifolia led to caulerpenyne and four new sulphated pyruvic acid derivatives. The work on the ascidian Polysyncraton sp. led to bolascidins A-D, four new metabolites which belong to the bolaamphiphile family, which, to the best of our knowledge, have no natural or synthetic equivalent. The first chemical study of the sponge Hexadella racovitzai led to two bromotyrosine derivatives, psammapline A and a new molecule, 4-O-sulfatocyclobispsammapline A. In addition, three new C1 crambescins, together with one crambescine A2 already reported, have been isolated from Crambe crambe. For all isolated marine invertebrate metabolites, the producer(s) remain(s) to be explored. The study of the marine fungus Stachybotrys chartarum, isolated from the sponge Aplysina cavernicola, led to satratoxin H and three stachybotrylactam derivatives already reported in the literature, and to 2,3-dihydrosatratoxin H and epi-chartarutine G, two new natural products. The different metabolites were evaluated for their antibacterial and cytotoxic properties. This work benefited from new analytical approaches (calculations of electronic circular dichroïsm spectra by TDDFT, molecular networking) and reinforced the interest of studies related to chemical ecology and search for new bioactive molecules to be valorized in different areas.