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.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/1160 |
| Date | 12 1900 |
| Creators | Martari, Marco |
| Contributors | Sanderson, R. D., University of Stellenbosch. Faculty of Science. Dept. of Chemistry and Polymer Science. |
| Publisher | Stellenbosch : University of Stellenbosch |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | 2808877 bytes, application/pdf |
| Rights | University of Stellenbosch |
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