THESIS
2014
iii leaves, iv-xv, 181 pages : illustrations (some color) ; 30 cm
Abstract
The marine environment is a plentiful source of bioactive natural products that can
assist us in the development of potential pharmaceutical agents. In the present study, we
screened 300 bacteria from the Red Sea for their antitumor and antibacterial activities. The
bioassay-guided analysis of the strain Streptomyces caelestis AW99C led to the discovery
of two diastereoisomers citreamicin ε A and B, together with their derivatives. The
antibacterial activities of all compounds were evaluated. Further cytotoxic mechanism
study revealed that citreamicin ε could cause DNA fragmentation and induce
caspase-3-dependent apoptosis in HeLa cells. Citreamicin ε greatly increased the
intracellular ROS concentration in HeLa cells, suggesting the possible involvement of ROS
in cell apoptosi...[
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The marine environment is a plentiful source of bioactive natural products that can
assist us in the development of potential pharmaceutical agents. In the present study, we
screened 300 bacteria from the Red Sea for their antitumor and antibacterial activities. The
bioassay-guided analysis of the strain Streptomyces caelestis AW99C led to the discovery
of two diastereoisomers citreamicin ε A and B, together with their derivatives. The
antibacterial activities of all compounds were evaluated. Further cytotoxic mechanism
study revealed that citreamicin ε could cause DNA fragmentation and induce
caspase-3-dependent apoptosis in HeLa cells. Citreamicin ε greatly increased the
intracellular ROS concentration in HeLa cells, suggesting the possible involvement of ROS
in cell apoptosis. Since citreamicin ε A and B had different apoptotic effects on PtK2 cells,
we performed an iTRAQ based quantitative proteomic analysis in order to reveal the
mechanism of cytotoxicity of citreamicin ε A and B in PtK2 cells. These significantly
changed proteins were analyzed for their protein-protein interaction, which revealed the
involvement of NF-κB pathway. Citreamicin ε A treatment induced rapid activation of the
NF-κB pathway, which might have promoted cell survival and resulted in lower toxicity.
Biosynthetic studies in the present thesis refered to two kinds of natural products: one was polycyclic xanthone antibiotics and another one was hybrid polyketide-terpenoid
halogenated meroterpenoids. The polycyclic xanthone antibiotics are synthesized by a type
II polyketide synthase (PKS) and then undergo various post-PKS modifications to produce
a variety of the polycyclic xanthone antibiotics. The entire 38-kb citreamicin (cit) gene
cluster was sequenced, and in silica analysis revealed 36 open reading frames (ORFs)
encoding type II polyketide synthases, regulators, transporters, and unique polyketide
tailoring enzymes. Regarding to the marinones which are hybrid polyketide-terpenoid
halogenated meroterpenoids, we adapted the transformation-associated recombination
(TAR) to clone the targeted 61-kb marinone biosynthesis (mrn) gene cluster. In addition, the
gene cluster unexpectedly harbored three vanadium-dependent haloperoxidases (VHPO)
genes that putatively brominated the naphthoquinone core via novel bacterial halogenation
biochemistry. To elucidate the divergent biosynthetic pathways of the marinones and the
function of the haloperoxidases, we performed a series of in vivo and in vitro experiments.
Our results confirmed that one of the VHPOs catalyzed the bromination of debromarinone
and neomarinone.
The finding of this thesis showed that 1) the Red Sea microbes are potent and
unexplored source of novel and bioactive compounds, 2) bioactive compounds indentified
by me showed antitumor and antibacterial activities, 3) comparative proteomic analysis
can be used to study the mode-of-action of stereoisomers in toxicology, and
4) the transformation-associated recombination cloning can be used in the biosynthetic
study of natural products from microbes.
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