Structure-function relationships of bolaamphiphilic peptides and peptide hybrids

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

Critical self-assembly concentration of bolaamphiphilic peptides andpeptide hybrids determined by fluorescence measurements

The original publication is available at http://www.scielo.org.zaCITATION: Martari, M. & Sanderson, R. D. 2013. Critical self-assembly concentration of bolaamphiphilic peptides andpeptide hybrids determined by fluorescence measurements. South African Journal of Chemistry, 61:47-52.The study of the self-assembly properties of peptides and proteins is important for the understanding of molecular recognition processes and for the rational design of functional biomaterials. Novel bolaamphiphilic peptides and peptide hybrids incorporating non-natural aminoacids were designed around a model lysine/leucine-rich peptide with the intention to study their selfassembly behaviour. Steady-state fluorescence measurements using pyrene as fluorescent probe were adapted to the determination of the critical self-assembly concentrations (CSACs) of these amphiphilic peptides and peptide hybrids. Different experimental conditions were studied. The morphology of the peptide aggregates was evaluated by scanning electron microscopy (SEM). Concentration andpHhave been revealed to play a key role in the control of the process. Peptides presented different three-dimensional supramolecular arrangements that were correlated with their aminoacid compositions (specifically considering the presence of tyrosine and proline) and CSAC values.Publisher's versio

CLUB Working Papers in Linguistics

CLUB WORKING PAPERS IN LINGUISTICS (CLUB-WPL) è una collana editoriale a cura del CLUB – Circolo Linguistico dell'Università di Bologna. La collana ospita contributi relativi alle iniziative del CLUB e dei suoi membri. I volumi, sottoposti a una procedura di peer-review, sono pubblicati esclusivamente online – sulla piattaforma AMS Acta dell'Università di Bologna – e sono liberamente accessibili

CLUB Working Papers in Linguistics

Con questo volume, la collana "CLUB Working Papers in Linguistics" giunge al suo terzo anno di vita, e presenta ancora una volta, con 18 contributi in formato open access, i risultati dello scambio di idee e di prospettive che si \ue8 sviluppato attraverso i vari incontri organizzati dal CLUB nell\u2019a.a. 2017\u201318. Il volume, che ospita anche i risultati del CLUB DAY su "Tipologia e diacronia: alla ricerca di sinergie", contiene saggi a firma di Fabio Ardolino, Silvia Ballar\ue8, Alessandra Barotto, Chiara Calderone, Sonia Cristofaro, Ilaria Fiorentini, Fernando Giacinti (vincitore del premio CLUB \u2018Una tesi in linguistica\u2019 per l\u2019anno 2018), Chiara Gianollo, Eugenio Goria, Nicola Grandi, Pierre Larriv\ue9e, Pauline Levillain, Edoardo Lombardi Vallauri, Elisabetta Magni, Yahis Martari, Francesca Masini, Simone Mattiola, Caterina Mauri, Marco Mazzoleni, Maria Napoli

Mutation Analysis of the Muscarinic Cholinergic Receptor Genes in Isolated Growth Hormone Deficiency Type IB

Background: Isolated GH deficiency (IGHD) is familial in 5–30% of patients. The most frequent form (IGHD-IB) has autosomal recessive inheritance, and it is known that it can be caused by mutations in the GHRH receptor (GHRHR) gene or in the GH gene. However, most forms of IGHD-IB have an unknown genetic cause. In normal subjects, muscarinic cholinergic stimulation causes an increase in pituitary GH release, whereas its blockade has the opposite effect, suggesting that a muscarinic acetylcholine receptor (mAchR) is involved in stimulating GH secretion. Five types of mAchR (M1–M5) exist. A transgenic mouse in which the function of the M3 receptor was selectively ablated in the central nervous system has isolated GH deficiency similar to animals with defective GHRH or GHRHR gene

Longevity in Untreated Congenital Growth Hormone Deficiency Due to a Homozygous Mutation in the GHRH Receptor Gene

Context: Reduced longevity observed in hypopituitarism has been attributed to GH deficiency (GHD). It is, however, unclear whether GHD or other confounding factors cause this early mortality