Thesis (PhD (Biochemistry))--University of Stellenbosch, 2010. / ENGLISH ABSTRACT:
The need for alternative or supplementary treatments due to the global problem of
microbial resistance towards conventional antimicrobials may be met by the
development of novel drugs based on antimicrobial peptides. The antimicrobial peptides
of interest to this study were the tyrocidines, cyclic decapeptides produced by Bacillus
aneurinolyticus. Although these antimicrobial peptides were the first natural antibiotic
to be discovered though a systematic search for antibacterial compounds, information
regarding their bioactivity, structure-activity relationships, determinants of bioactivity
and mode of action is limited. The aim of this study was to investigate the antibacterial
and antiplasmodial activity, as well as to identify determinants of bioactivity
modulation, of the natural tyrocidine library.
The study indicated that the tyrocidines exhibit significant activity toward
Gram-positive bacteria, notably Listeria monocytogenes, and the intraerythocytic
parasite, Plasmodium falciparum. Both the antilisterial and antiplasmodial activity was
found to be highly dependent on peptide identity and self-assembly. The antilisterial
activity of the tyrocidines was shown to be associated with increased self-assembly
within a membrane-like environment, which suggested that formation of lytic
complexes within the bacterial membrane may play a crucial role in tyrocidine activity.
In contrast to the observations for antilisterial activity, the antiplasmodial activity of the
tyrocidines was shown to be associated with reduced self-assembly within a
membrane-like environment, which suggested that the antiplasmodial activity of the
tyrocidines is mediated by a mechanism other than the formation of lytic complexes
within the target cell membrane.
In addition to the influence of peptide identity and self-assembly, the bioactivity of the
tyrocidines was found to be highly sensitive to environmental conditions, notably the
presence of calcium. The antilisterial activity, as well as the mode of action, of the
tyrocidines was also found to be highly sensitive to tyrocidine-Ca2+ complexation and
the concomitant induction of higher-order structures. Tyrocidine-Ca2+ complexation
was shown to greatly enhance antilisterial activity and change the mechanism of action
from a predominantly membranolytic to an alternative, non-lytic mode of action. The results of this investigation suggest that the alternative mode of tyrocidine activity
may be related to complexation with Ca2+. It is hypothesised that such complexation
may either (1) promote tyrocidine-DNA complexation, and thus inhibition of
transcription and/or replication; or (2) interfere with Ca2+ homeostasis, and thus
influence vital cell functions.
Overall, it may be hypothesised that tyrocidine activity and mode of action is modulated
by a critical play-off between self-assembly, cation-complexation and
membrane-interaction. As these modulators of activity are highly dependent on
tyrocidine sequence/structure, the wide variety of tyrocidines found in the natural
complex may allow for optimal interaction with and activity toward a variety of
microbes. / AFRIKAANSE OPSOMMING:
Die universele probleem van mikrobiese weerstand teen konvensionele antimikrobiese
middels en die wêreld-wye noodsaaklikheid vir alternatiewe of bykomende behandeling
mag deur die ontwikkeling van nuwe middels, gebasseer op antimikrobiese peptiede,
vervul word. Die antimikrobiese peptiede van belang tot hierdie studie is die tirosidiene,
sikliese dekapeptiede wat deur Bacillus aneurinolyticus geproduseer word. Informasie
ten opsigte van die tirosidiene se bioaktiwiteit, struktuur-funksieverwantskap,
determinante van bio-aktiwiteit en meganisme van aksie was beperk, alhoewel hierdie
peptiede die eerste antimikrobiese peptiede was wat ontdek is deur ‘n sistematiese
soektog vir antimikrobiese middels. Die doelwit van hierdie studie was die ondersoek
van antibakteriële and antiplasmodiese aktiwiteit, sowel as om die determinante van
bio-aktiwiteit modulering van die natuurlike tirosidienbiblioteek te ondersoek.
Hierdie studie het getoon dat die tirosidiene merkwaardige aktiwiteit teenoor
Gram-positiewe bakterië, in besonder Listeria monocytogenes het, asook teenoor die
intra-eritrositiese parasiet, Plasmodium falciparum. Daar is bevind dat beide die antilisteriese
en antiplasmodiese aktiwiteite hoogs afhanklik is van peptiedidentiteit en
self-verpakking. Daar is gewys dat die antilisteriese aktiwiteit van die tirosidiene
geassosieer is met verhoogde self-verpakking in ’n membraanagtige omgewing, wat ’n
aanduiding is dat die vorming van litiese komplekse in die bakteriële membraan ’n
kritiese rol in tirosidienaktiwiteit speel. Kontrasterend tot die waarnemings van
antilisteriese aktiwiteit, is getoon dat die antiplasmodiese aktiwiteit van die tirosidiene
geassosieer is met verlaagde self-verpakking in ’n membraanagtige omgewing. Dis ’n
aanduiding dat die antiplasmodiese aktiwiteit van die tirosidiene gemediëer word deur
‘n ander meganisme en nie die vorming van litiese komplekse in die teikenselmembraan
nie.
Bykomend tot die invloed van peptiedidentiteit en self-verpakking, is daar bevind dat
die bioaktiwiteit van die tirosidiene hoogs sensitief is vir die omgewing, in besonder die
teenwoordigheid van kalsium. Daar is ook bevind dat die antilisteriese aktiwiteit, sowel
as die meganisme van aksie, van tirosidiene hoogs sensitief is vir tirosidien-Ca2+
kompleksvorming en die gevolglike induksie van of hoër-orde strukture. Daar is gewys dat tirosidien-Ca2+ kompleksvorming die antilisteriese aktiwiteit drasties verhoog en dat
die meganisme van aksie verander van ’n oorwegende membranolitiese meganisme na
’n alternatiewe nie-litiese meganisme van aksie.
Die resultate van hierdie ondersoek het aangedui dat die alternatiewe meganisme van
aksie van tirosidienaktiwiteit moontlik verband kan hou met kompleksvorming met
Ca2+. Die hipotese is dat sodanige kompleksvorming moontlik of (1) tirosidien-DNA
komplekvorming aanmoedig, en dus transkripsie en/of replikasie inhibibeer of (2) met
Ca2+ homeostase inmeng, en sodoende lewensnoodsaaklike selfunksies beïnvloed.
Die algemene hipotese is dat tirosidienaktiwiteit en meganisme van aksie deur ’n
kritiese spel tussen self-verpakking, katioonkompleksvorming en membraaninteraksie
gemoduleer word. Die wye verskeidenheid van tirosidiene, wat in die natuurlike
kompleks gevind word, kan moontlik toelaat vir die optimale interaksie met, en
aktiwiteit teenoor ’n verskeidenheid van mikrobes, aangesien die aktiwiteitmoduleerders hoogs afhanklik is van tirosidien struktuur/volgorde.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/4001 |
| Date | 03 1900 |
| Creators | Spathelf, Barbara Marianne |
| Contributors | Rautenbach, M., University of Stellenbosch. Faculty of Science. Dept. of Biochemistry. |
| Publisher | Stellenbosch : University of Stellenbosch |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | English |
| Type | Thesis |
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